4.3: Glance at Genre- Conflict, Detail, and Revelation
By the end of this section, you will be able to:
- Identify common formats and design features used to develop a personal narrative or memoir.
- Show that genre conventions are shaped by purpose, culture, and expectation.
In personal writing genres, you share experiences of your life, centering them on a specific theme or memory. Unlike an autobiography, which typically extends across an entire lifetime or at least a number of years, memoirs and personal narratives are shaped by a narrower focus, with more specific storytelling surrounding a time period or an event. When writing in these personal genres, authors seek to make an emotional connection with their audience to relate an experience, emotion, or lesson learned.
Characteristics of Memoirs and Personal Narratives
One way to approach a memoir or personal narrative is to think of it as a written series of photographs—snapshots of a period of time, moment, or sequence of events connected by a theme. In fact, writing prose snapshots is analogous to constructing and arranging a photo album composed of separate images. Photo albums, when carefully assembled from informational snapshots, tell stories with clear beginnings, middles, and endings. However, they show a lot of white space between one picture and the next, with few transitions explaining how the photographer got from one scene to another
In other words, while photo albums tell stories, they do so piecemeal, requiring viewers to fill in or imagine what happens between shots. You also might think of snapshots as individual slides in a slideshow or pictures in an exhibition—each the work of the same maker, each a different view, all connected by some logic, the whole presenting a story.
Written snapshots function in the same way as visual snapshots, each connected to the next by white space. Sometimes written snapshots can function as a series of complete and independent paragraphs, each an entire thought, without obvious connections or transitions to the preceding or following paragraph. White space between one snapshot and another gives readers breathing space, allowing them time to digest one thought before continuing to the next. It also exercises readers’ imaginations; as they participate in constructing logic that offers textual meaning, the readers themselves make connections and construct meaning. At other times, snapshots flow more directly, one after another, through chronological, circular, parallel, or other structures to move from event to event.
The secret to using snapshots successfully in your writing is to place them carefully in an order that conveys a theme and creates an unbreakable thread. And as with visual snapshots, writers must carefully choose which moments to include—and which to omit. Because they tell stories, memoirs and personal narratives share aspects of the fictional narrative genre. In writing them, you will use crafting tools to tell a vivid and purposeful story that takes into consideration your personal experience and the needs of your reader.
The Story Teller's Dilemma: Clarity of Action
How you construct your story is as important as the story you choose to tell. Deciding on the most effective way the way to tell the story—that is, deciding what framework to use—helps you develop clarity of action to lead readers to the theme or message you seek to develop. Various components work together to bring clarity, but most often in a memoir or personal narrative, clarity comes from plot and character development. Narratives often follow a general structure called an arc to develop characters and plot and build the emotional impact of a story. Look at Figure \(4.4\) for an idea of what a story arc looks like.
Figure \(4.4\) Narrative arc (CC BY 4.0; Rice University & OpenStax)
This arc, also called a narrative arc or a plot triangle , is composed of the following elements:
- The exposition sets up the narrative. It introduces characters and setting and establishes the primary conflict of the story, allowing readers to learn the who, what, when, where, and why of the events that will unfold.
- Next, the rising action fully develops the conflict. This developed series of events, the longest part of the narrative, produces increasing tension that engages readers.
- The climax is the turning point of the narrative, in which the story reaches its highest point of tension and conflict. It is the moment when some kind of action must be taken.
- In the falling action , the conflict begins to be resolved, and the tension lessens.
- Finally, during the resolution , the conflict is resolved, and the narrative ends. In memoirs and personal narratives, the resolution often includes or precedes a reflection that examines the broader implications of the theme or lessons learned. Of course, as in real life, conflicts are not always resolved, but the narrator can still reflect on the outcome of the situation.
Although many narratives and memoirs follow this plot-driven arc, narratives can also focus on character arcs. Character-driven narratives explore an individual, most often the narrator, and their development. The stories focus on creating an emotional connection between the character and the reader. Both plot and character arcs may be, and often are, present in memoirs and personal narratives.
Regardless of whether the focus is on plot or characters, conflict is synonymous with the reason for telling your story—it is the driving force. Conflict is often the main challenge faced by a character, and it urges the story along by engaging readers through tension and encouraging them to keep reading. Without conflict, your memoir or personal narrative will lack an overall theme. The major conflict is the undercurrent that drives each scene and is often developed by an inciting incident . Introduced in the exposition and developed in the rising action, this incident sets the mood of the story and engages readers. After the story’s climax, where the conflict reaches its peak, the tension gradually resolves during the falling action and moves toward resolution, during which you can explicitly or implicitly explore the theme that ties the story elements together. Sometimes the resolution is accompanied by a revelation , in which the narrator or reader understands something about the bigger picture, such as a lesson learned from the events recounted or knowledge about the general human condition. Of course, each scene or section should have its own conflict, connected in some way to the overarching message of your writing. As you write, ask yourself, What’s the conflict? By identifying the conflict explicitly, you will ensure that it remains central to your narrative.
Two important aspects of plot structure regarding time in a memoir are chronos and kairos . Chronos is the sequence of events told according to their order. This order is often chronological and linear, but not always—it can be interrupted, fragmented, circular, or otherwise out of sequence and can sometimes include flashbacks. Chronos develops themes by the telling of events. Kairos , on the other hand, is the Greek concept of timeliness. Events told through the lens of kairos are often transcendental, an argument that is made at the right time, often rooted in a cultural moment or movement.
Other important aspects of personal writing overlap with the narrative genre. Both reader engagement and plot rely largely on vivid details and sensory descriptions to move readers through the story. For more information on narrative elements that may enhance your personal narrative or memoir, revisit Literacy Narrative: Building Bridges, Bridging Gaps.
Key Terms for Memoir or Personal Narrative Writing
- Anecdotes: a short, interesting story or event told to demonstrate a point or amuse the audience.
- Bias: the inclusion or exclusion of certain events and facts, the decisions about word choice, and the consistency of tone. All work together to convey a particular feeling or attitude. Bias comes from a specific stance or worldview and can limit a text, particularly if that bias is left unexamined.
- Characters: fictional people (or other beings) created in a work of literature. The narrator of a memoir or personal narrative is the nonfiction equivalent of the main character.
- Climax: the point of highest level of interest and emotional response in a narrative.
- Conclusion: in narrative writing, the resolution. It is the point at which the narrator has reached a decision.
- Conflict: the major challenge that the main character faces.
- Doubling: a mirroring of events, objects, characters, or concepts in a memoir.
- Exposition: the beginning section of a narrative that introduces the characters, setting, and plot.
- Falling action: the section of the plot after the climax in which tension from the main conflict is decreased and the narrative moves toward the conclusion, or resolution.
- Flashback: a scene that interrupts the chronological order of the main narrative to return to a scene from an earlier time.
- Foreshadowing: hints of what is to come in the text.
- Mood: the atmosphere of the text, often achieved through details, description, and setting.
- Plot: the events that make up a narrative or story.
- Point of view: the perspective from which a narrative is told. Memoirs and personal narratives usually use the first-person point of view, or tell the story through the eyes of the narrator.
- Resolution: the point at which a story’s conflict is settled; the conclusion of a narrative.
- Revelation: a discovery about a person, event, or idea that shapes the plot.
- Rising action: a series of events in the plot in which tension surrounding the major conflict increases and the plot moves toward its climax.
- Setting: when and where a narrative occurs. Setting is revealed through narration and details.
- Theme: the underlying idea that reveals the author’s message about a narrative.
- Vivid details: sensory language and detailed descriptions that help readers gain a deeper and fuller understanding of ideas and events in the narrative.
- Voice: the combination of vocabulary, tone, sentence structure, dialogue, and other details that make a text authentic and engaging. Voice is the “identity” or “personality” of the writer and includes the specific English variety used by the narrator and characters.


3.2.4: Cho Se-hui (1942- )
Selected Stories Korean Postcolonialism / Contemporary Literature Cho Se-hui (surname: Cho) is a South Korean writer of the so-called "hangeul [the Korean alphabet] generation," which refers to the first generation to be educated entirely in Korean, unlike the previous generations who received Japanese colonial education or studied Chinese primarily. Cho is most well-known for his novel A Little Ball Launched by a Dwarf (1978), which was initially published as a "linked novel"—a series of related short stories in several Korean magazines. "Knifeblade," "A Little Ball Launched by a Dwarf," and "The Möbius Strip" are parts of this novel, which is noted for its sharp allegorical social criticism of South Korea in the 1970s. Written during the Park Chung-hee authoritarian regime (1961-1979), when South Korea was undergoing rapid industrialization, the novel focuses on the forced redevelopment of Seoul in the 1970s and asks at what human cost the economic development has taken place. Written in concise and accessible language and with the use of irony, the novel brings social contradictions, labor issues under capitalism, and the relationship between the haves and have-nots to the fore. The English translation of the novel, titled The Dwarf , by Bruce and Ju-Chan Fulton was published in 2006. Cho's other works include Time Travel (1983) and Root of Silence (1985). Consider while reading:
- What aspects of South Korean society are being satirized by Cho?
- How is irony employed in these stories? Provide examples.
- What is the significance of these short stories' titles?
- Do some research on the history of Korea, and explore how some of the legacies of Japanese colonialism in Korea might be related to these short stories.
Written by Kyounghye Kwon


1.1: Water Cycle
Learning Objectives
After reading this section, you should be able to:
- Identify processes in the water cycle that influence the water supply
- Analyze situations in terms of precipitation, condensation, evaporation, and transpiration
- Evaluate yearly data for evapotranspiration
You've probably heard on the news the statewide water supply in California described in terms of snowpack and rainfall. These are critical measures of how much water is available for human use. Water managers in California follow both snowpack measurements and rainfall measurements closely. In the diagram above from the United States Geological Survey (USGS), you can see that precipitation is shown as rainfall and snow.
Precipitation also includes something you don’t see too often in California: hail and sleet. All forms of water that fall from the sky, including rain, snow, hail and sleet, are forms of precipitation. Rainfall can provide much needed water in the ground by the process of infiltration, the process by which water seeps into the ground and eventually recharges our groundwater, water stored in the ground. Using the diagram above, you can also trace the flow of groundwater in some cases to both rivers, lakes, and even to the ocean.
Above ground, rainfall can also be stored on earth’s surface in lakes. Precipitation in California also provides snow for snowpack, which can be our largest area of storage of water in the winter. In fact, it is common for water managers to view the snowpack in the Sierra as a reservoir; it is simply a seasonal reservoir that melts in the spring. Snowmelt runoff from the snowpack can fill streams and lakes. Runoff from rainfall is also captured in streams and lakes and can serve to recharge aquifers through percolation and infiltration.
The diagram also shows the key process of evaporation, or the process through which liquid water turns into a gas. Evaporation occurs over bodies of water like the ocean or lakes, but also over the land. If you own a home with a pool, you have probably noticed that if you leave the pool without a cover in the summer, you have to add water more often in the summer than in the winter. This is because the rate of evaporation is higher in the summer than the winter, often dramatically so.
Transpiration is the process through which plants lose water. This may seem like an inconsequential process, but it is the entire process that drives irrigation. In the diagram below you can follow the transpiration process. In Step 1, plants bring in moisture from the soil with their roots. Then, in Step 2, water travels up through the plant. In Step 3, water leaves through their pores or stomates (plural of stomata) and enters the atmosphere again.
The combination of water loss from liquid water in the ground evaporating and water in plants losing water is called evapotranspiration (ET). Evapotranspiration is frequently used by scientists as a measurement of the plant water needs. If a location has a high ET, then the plant needs are greater than a location with a low ET.
Most locations in the state of California are close to a weather station that is part of the California Irrigation Management Information System (CIMIS) network. A CIMIS station measures a variety of variables in order to calculate the ET including: temperature, solar radiation, humidity, wind speed, and wind direction. With a complicated formula, the CIMIS station will calculate the ET for a specific geographic location. This information is frequently used to make a water budget, an estimate of how much water a location should use, including outdoor watering based on the amount of landscaping, the type of landscaping and the evapotranspiration.
The monthly inches of ET data in the table for Santa Clarita reflects the water needs in terms of inches of water that grass would need to receive per month. You can see that the highest needs are in summer, primarily July and August, with needs decreasing rapidly in September through December. Even though September is typically almost as hot as August, the days are shorter and the sun is less intense so the evapotranspiration is less in September than August. This means the plant needs for irrigation are significantly less.
CIMIS Station #204 Santa Clarita - 2016 Evapotranspiration Data
|
Months |
Jan |
Feb |
Mar |
Apr |
May |
Jun |
Jul |
Aug |
Sep |
Oct |
Nov |
Dec |
|
Monthly ET (in) |
1.97 |
4.80 |
4.51 |
5.24 |
5.77 |
7.77 |
8.99 |
8.13 |
6.30 |
4.68 |
3.40 |
2.69 |
Knowledge of evapotranspiration has a practical application - in Southern California, many people find they can turn off their irrigation systems for grass in normal years from November through February and that their plants’ water needs will be met with rainfall alone.
The opposite process of evaporation is condensation, the process through which water as a gas turns into liquid water again. Condensation is the process through which clouds form. This is also the process through which water beads on the outside of a glass of iced tea on a humid day or on a mirror in a steamy bathroom. In terms of the water supply picture, condensation is the key process that can lead to precipitation, so clouds are carefully monitored by scientists.
What makes the water cycle work? In short, gravity and the sun. Notice that there is always groundwater flow and streamflow from a higher altitude to a lower altitude. Gravity drives this flow of water in the water cycle, and is also a key force in most water distribution systems. If you’re not using gravity, you’re going to need a pump to go from a lower elevation to a higher elevation. Systems are generally designed to avoid pumps if possible and use the force of gravity. The sun drives the water cycle in a different way, by heating the water and causing it to evaporate. It also melts the snowpack in the mountains while gravity causes the water to flow into streams, rivers and lakes.
In Section 1.1, you've learned the basic processes in the water cycle: precipitation, infiltration, evaporation, transpiration, and condensation. Next, you'll see how these processes create surface and groundwater supplies and how these supplies are used for water management.
Try It!
Describe a process at work in each scenario and how the process works:
- Analyze the effect on the water cycle if California had no precipitation for a year.
- You hang a wet swimsuit out on the balcony to dry overnight. What process is at work? How is water changing states?
- Using the CIMIS web site (www.cimis.water.ca.gov/), register for an account (free). Then under the Data tab, find the nearest CIMIS station to your home create a monthly web report for 12 consecutive months, which should include ET. When should you water the most? When should you water the least?
Key Terms
Condensation—The process through which water as a gas turns into liquid water again; opposite of evaporation
Evaporation—The process through which liquid water turns into a gas; opposite of condensation
Evapotranspiration—The combination of water loss from liquid water in the ground evaporating and water in plants losing water
Groundwater—Water stored in the ground
Infiltration—The process through which water seeps into the ground
Precipitation—All forms of water that fall from the sky, including rain, snow, hail, and sleet
Transpiration—The process through which plants lose water
Water budget—An estimate of how much water a location should use, including outdoor watering based on the amount of landscaping, the type of landscaping and the evapotranspiration


2.16: Chemistry
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- Susan Rahman, Prateek Sunder, and Dahmitra Jackson
- CC ECHO
Chemistry is the branch of science that deals with matter and how it changes. This includes the identification of substances composed of matter, the investigation of atomic and molecular properties and the ways in which they interact, combine, and change, as well as the use of these processes to form new substances. The field of Chemistry, as with the majority of the Science, Technology, Engineering and Math (STEM) fields, is permeated with an historical legacy of structural racism.
In 2020, a conversation took place during #BlackInChem week about the lack of representation in the field and micro aggressions felt by Black Chemistry students. Looking at ways to address this involved surveying students of color in the field of chemistry about their experiences and what steps they perceived would be helpful. Many students cited a need for effective mentorship not just from faculty, but peer mentorship as well. They also stressed the need for greater financial support to allow them to devote time studying rather than working side jobs in order to survive. When students come from families that do not have prior experience with higher education, perhaps they are first generation college students, they also need help navigating the educational system (Remmel, 2020 12:15). Mentorship can come from internships in industry or research with faculty members in their academic labs. There has been funding put in place to target diversity in STEM via both of these types of Mentorships. Also knowing about scientists of color is empowering for students. Teachers have an opportunity to highlight those individuals as a part of their curriculum. The Marberger STEM Center at Lawrence Technological University ( https://www.ltu.edu/stem-center/ ) is a great resource for STEM faculty wishing to make their teaching more inclusive. It is important to note that the responsibility of diversifying the field cannot just fall on the Black scientists but rather a united effort on the part of all scientifically minded educators and researchers.
What will become clear as you read through this primer is that representation matters. It is important both for students of color and women to see themselves in their fields of study, so hiring practices in chemistry should reflect that goal. Additionally, for those faculty who do not represent the demographic of their students, it is important to highlight prominent thinkers in their field that are not just white men. In an effort to learn more about the history of Black Chemists here we highlight the first African American to earn a PhD in Chemistry.
Born in Louisville, Kentucky in 1884, Saint Elmo Brady became the first African American to earn a Ph.D. in the field of chemistry when he completed his graduate studies at the University of Illinois in 1916. Brady studied chemistry at Fisk University in Nashville, Tennessee and earned hisB.S. degree in 1908. After graduation he accepted a faculty position at Tuskegee Normal and Industrial Institute (now known as Tuskegee University) and was mentored by George Washington Carver and Booker T. Washington,President of Tuskegee. He began graduate studies in the Department of Chemistry at the University of Illinois in 1912, earning his M.S. degree in 1914 and completing his doctorate in 1916.
As a graduate student at Illinois, Brady’s research focused on the characterization of organic acids. Brady eventually published three abstracts focused on his graduate work in Science, a prestigious peer-reviewed journal.After completing his doctorate, Brady returned to Tuskegee and continued teaching until 1920. He also served as chair and faculty of the Department of Chemistry at Howard University and Fisk University. (Collins, 2007).
In the summer of 2020, the president of the American Chemical Society (ACS) issued a statement in response to the killing of George Floyd. As one of the world's largest scientific organizations they felt the use of their platform to connect the field of chemistry to social justice and structural racism was important (Echegoyen, 2020). There is an acknowledgement on the part of the American Chemical Society (ACS) that their publications do not represent enough diversity of voice and they released this statement denouncing racial violence. They, like other academic organizations, have committed to a number of first action steps including a commitment to diversity of journal contributors as a part of the evaluation of the Editor in Chief(Advancing ACS’, nd). Placing the task of ensuring diverse representation in the publication they edit as a part of their job performance ensures accountability and the outcome is easily measurable. In order to do so, it will be important to circle back to see how this equity work shifts the demographics of the field.


5.13: Electric Potential Field due to a Continuous Distribution of Charge
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The electrostatic potential field at \({\bf r}\) associated with \(N\) charged particles is
\[V({\bf r}) = \frac{1}{4\pi\epsilon} \sum_{n=1}^N { \frac{q_n}{\left|{\bf r}-{\bf r}_n\right|} } \label{m0065_eCountable} \]
where \(q_n\) and \({\bf r_n}\) are the charge and position of the \(n^{\mbox{th}}\) particle. However, it is more common to have a continuous distribution of charge as opposed to a countable number of charged particles. We now consider how to compute \(V({\bf r})\) three types of these commonly-encountered distributions. Before beginning, it’s worth noting that the methods will be essentially the same, from a mathematical viewpoint, as those developed in Section 5.4; therefore, a review of that section may be helpful before attempting this section.
Continuous Distribution of Charge Along a Curve
Consider a continuous distribution of charge along a curve \(\mathcal{C}\). The curve can be divided into short segments of length \(\Delta l\). Then, the charge associated with the \(n^{\mbox{th}}\) segment, located at \({\bf r}_n\), is
\[q_n = \rho_l({\bf r}_n)~\Delta l \nonumber \]
where \(\rho_l\) is the line charge density (units of C/m) at \({\bf r}_n\). Substituting this expression into Equation \ref{m0065_eCountable}, we obtain
\[{\bf V}({\bf r}) = \frac{1}{4\pi\epsilon} \sum_{n=1}^{N} { \frac{\rho_l({\bf r}_n)}{\left|{\bf r}-{\bf r}_n\right|} \Delta l} \nonumber \]
Taking the limit as \(\Delta l\to 0\) yields:
\[V({\bf r}) = \frac{1}{4\pi\epsilon} \int_{\mathcal C} { \frac{\rho_l(l)}{\left|{\bf r}-{\bf r}'\right|} dl} \label{m0065_eLineCharge} \]
where \({\bf r}'\) represents the varying position along \({\mathcal C}\) with integration along the length \(l\).
Continuous Distribution of Charge Over a Surface
Consider a continuous distribution of charge over a surface \(\mathcal{S}\). The surface can be divided into small patches having area \(\Delta s\). Then, the charge associated with the \(n^{\mbox{th}}\) patch, located at \({\bf r}_n\), is
\[q_n = \rho_s({\bf r}_n)~\Delta s \nonumber \]
where \(\rho_s\) is surface charge density (units of C/m\(^2\)) at \({\bf r}_n\). Substituting this expression into Equation \ref{m0065_eCountable}, we obtain
\[V({\bf r}) = \frac{1}{4\pi\epsilon} \sum_{n=1}^{N} { \frac{\rho_s({\bf r}_n)}{\left|{\bf r}-{\bf r}_n\right|}~\Delta s} \nonumber \]
Taking the limit as \(\Delta s\to 0\) yields:
\[V({\bf r}) = \frac{1}{4\pi\epsilon} \int_{\mathcal S} { \frac{\rho_s({\bf r}')}{\left|{\bf r}-{\bf r}'\right|}~ds} \label{m0065_eSurfCharge} \]
where \({\bf r}'\) represents the varying position over \({\mathcal S}\) with integration.
Continuous Distribution of Charge in a Volume
Consider a continuous distribution of charge within a volume \(\mathcal{V}\). The volume can be divided into small cells (volume elements) having area \(\Delta v\). Then, the charge associated with the \(n^{\mbox{th}}\) cell, located at \({\bf r}_n\), is
\[q_n = \rho_v({\bf r}_n)~\Delta v \nonumber \]
where \(\rho_v\) is the volume charge density (units of C/m\(^3\)) at \({\bf r}_n\). Substituting this expression into Equation \ref{m0065_eCountable}, we obtain
\[V({\bf r}) = \frac{1}{4\pi\epsilon} \sum_{n=1}^{N} { \frac{\rho_v({\bf r}_n)}{\left|{\bf r}-{\bf r}_n\right|}~\Delta v} \nonumber \]
Taking the limit as \(\Delta v\to 0\) yields:
\[V({\bf r}) = \frac{1}{4\pi\epsilon} \int_{\mathcal V} { \frac{\rho_v({\bf r}')}{\left|{\bf r}-{\bf r}'\right|}~dv} \nonumber \]
where \({\bf r}'\) represents the varying position over \({\mathcal V}\) with integration.


4.3: Covalent Bonding
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Learning Objectives
By the end of this section, you will be able to:
- Describe the formation of covalent bonds
- Define electronegativity and assess the polarity of covalent bonds
Ionic bonding results from the electrostatic attraction of oppositely charged ions that are typically produced by the transfer of electrons between metallic and nonmetallic atoms. A different type of bonding results from the mutual attraction of atoms for a “shared” pair of electrons. Such bonds are called covalent bonds . Covalent bonds are formed between two atoms when both have similar tendencies to attract electrons to themselves (i.e., when both atoms have identical or fairly similar ionization energies and electron affinities). For example, two hydrogen atoms bond covalently to form an H 2 molecule; each hydrogen atom in the H 2 molecule has two electrons stabilizing it, giving each atom the same number of valence electrons as the noble gas He.
Compounds that contain covalent bonds exhibit different physical properties than ionic compounds. Because the attraction between molecules, which are electrically neutral, is weaker than that between electrically charged ions, covalent compounds generally have much lower melting and boiling points than ionic compounds. In fact, many covalent compounds are liquids or gases at room temperature, and, in their solid states, they are typically much softer than ionic solids. Furthermore, whereas ionic compounds are good conductors of electricity when dissolved in water, most covalent compounds are insoluble in water; since they are electrically neutral, they are poor conductors of electricity in any state.
Formation of Covalent Bonds
Nonmetal atoms frequently form covalent bonds with other nonmetal atoms. For example, the hydrogen molecule, H 2 , contains a covalent bond between its two hydrogen atoms. Figure \(\PageIndex{1}\) illustrates why this bond is formed. Starting on the far right, we have two separate hydrogen atoms with a particular potential energy, indicated by the red line. Along the x -axis is the distance between the two atoms. As the two atoms approach each other (moving left along the x -axis), their valence orbitals (1 s ) begin to overlap. The single electrons on each hydrogen atom then interact with both atomic nuclei, occupying the space around both atoms. The strong attraction of each shared electron to both nuclei stabilizes the system, and the potential energy decreases as the bond distance decreases. If the atoms continue to approach each other, the positive charges in the two nuclei begin to repel each other, and the potential energy increases. The bond length is determined by the distance at which the lowest potential energy is achieved.
It is essential to remember that energy must be added to break chemical bonds (an endothermic process), whereas forming chemical bonds releases energy (an exothermic process). In the case of H 2 , the covalent bond is very strong; a large amount of energy, 436 kJ, must be added to break the bonds in one mole of hydrogen molecules and cause the atoms to separate:
\[\ce{H2(g) -> 2 H(g)} \quad \Delta H=436 \,\text{kJ} \nonumber \]
Conversely, the same amount of energy is released when one mole of H 2 molecules forms from two moles of H atoms:
\[\ce{2 H(g) -> H2(g)} \quad \Delta H=-436\,\text{kJ} \nonumber \]
Pure vs. Polar Covalent Bonds
If the atoms that form a covalent bond are identical, as in H 2 , Cl 2 , and other diatomic molecules, then the electrons in the bond must be shared equally. We refer to this as a pure covalent bond . Electrons shared in pure covalent bonds have an equal probability of being near each nucleus.
In the case of Cl 2 , each atom starts off with seven valence electrons, and each Cl shares one electron with the other, forming one covalent bond:
\[\ce{Cl + Cl -> Cl2} \nonumber \]
The total number of electrons around each individual atom consists of six nonbonding electrons and two shared (i.e., bonding) electrons for eight total electrons, matching the number of valence electrons in the noble gas argon. Since the bonding atoms are identical, Cl 2 also features a pure covalent bond.
When the atoms linked by a covalent bond are different, the bonding electrons are shared, but no longer equally. Instead, the bonding electrons are more attracted to one atom than the other, giving rise to a shift of electron density toward that atom. This unequal distribution of electrons is known as a polar covalent bond , characterized by a partial positive charge on one atom and a partial negative charge on the other. The atom that attracts the electrons more strongly acquires the partial negative charge and vice versa. For example, the electrons in the H–Cl bond of a hydrogen chloride molecule spend more time near the chlorine atom than near the hydrogen atom. Thus, in an HCl molecule, the chlorine atom carries a partial negative charge and the hydrogen atom has a partial positive charge. Figure \(\PageIndex{2}\) shows the distribution of electrons in the H–Cl bond. Note that the shaded area around Cl is much larger than it is around H. Compare this to Figure \(\PageIndex{1}\), which shows the even distribution of electrons in the H 2 nonpolar bond.
We sometimes designate the positive and negative atoms in a polar covalent bond using a lowercase Greek letter “delta,” δ, with a plus sign or minus sign to indicate whether the atom has a partial positive charge (δ+) or a partial negative charge (δ–). This symbolism is shown for the H–Cl molecule in Figure \(\PageIndex{2}\).
Electronegativity
Whether a bond is nonpolar or polar covalent is determined by a property of the bonding atoms called electronegativity . Electronegativity is a measure of the tendency of an atom to attract electrons (or electron density) towards itself. It determines how the shared electrons are distributed between the two atoms in a bond. The more strongly an atom attracts the electrons in its bonds, the larger its electronegativity. Electrons in a polar covalent bond are shifted toward the more electronegative atom; thus, the more electronegative atom is the one with the partial negative charge. The greater the difference in electronegativity, the more polarized the electron distribution and the larger the partial charges of the atoms.
Figure \(\PageIndex{3}\) shows the electronegativity values of the elements as proposed by one of the most famous chemists of the twentieth century: Linus Pauling (Figure \(\PageIndex{4}\)). In general, electronegativity increases from left to right across a period in the periodic table and decreases down a group. Thus, the nonmetals, which lie in the upper right, tend to have the highest electronegativities, with fluorine the most electronegative element of all (EN = 4.0). Metals tend to be less electronegative elements, and the group 1 metals have the lowest electronegativities. Note that noble gases are excluded from this figure because these atoms usually do not share electrons with others atoms since they have a full valence shell. (While noble gas compounds such as XeO 2 do exist, they can only be formed under extreme conditions, and thus they do not fit neatly into the general model of electronegativity.)
Electronegativity versus Electron Affinity
We must be careful not to confuse electronegativity and electron affinity. The electron affinity of an element is a measurable physical quantity, namely, the energy released or absorbed when an isolated gas-phase atom acquires an electron, measured in kJ/mol. Electronegativity, on the other hand, describes how tightly an atom attracts electrons in a bond. It is a dimensionless quantity that is calculated, not measured. Pauling derived the first electronegativity values by comparing the amounts of energy required to break different types of bonds. He chose an arbitrary relative scale ranging from 0 to 4.
Portrait of a Chemist: Linus Pauling
Linus Pauling , shown in Figure \(\PageIndex{4}\), is the only person to have received two unshared (individual) Nobel Prizes: one for chemistry in 1954 for his work on the nature of chemical bonds and one for peace in 1962 for his opposition to weapons of mass destruction. He developed many of the theories and concepts that are foundational to our current understanding of chemistry, including electronegativity and resonance structures.
Pauling also contributed to many other fields besides chemistry. His research on sickle cell anemia revealed the cause of the disease—the presence of a genetically inherited abnormal protein in the blood—and paved the way for the field of molecular genetics. His work was also pivotal in curbing the testing of nuclear weapons; he proved that radioactive fallout from nuclear testing posed a public health risk.
Electronegativity and Bond Type
The absolute value of the difference in electronegativity (ΔEN) of two bonded atoms provides a rough measure of the polarity to be expected in the bond and, thus, the bond type. When the difference is very small or zero, the bond is covalent and nonpolar. When it is large, the bond is polar covalent or ionic. The absolute values of the electronegativity differences between the atoms in the bonds H–H, H–Cl, and Na–Cl are 0 (nonpolar), 0.9 (polar covalent), and 2.1 (ionic), respectively. The degree to which electrons are shared between atoms varies from completely equal (pure covalent bonding) to not at all (ionic bonding). Figure \(\PageIndex{5}\) shows the relationship between electronegativity difference and bond type.
A rough approximation of the electronegativity differences associated with covalent, polar covalent, and ionic bonds is shown in Figure \(\PageIndex{5}\). This table is just a general guide, however, with many exceptions. For example, the H and F atoms in HF have an electronegativity difference of 1.9, and the N and H atoms in NH 3 a difference of 0.9, yet both of these compounds form bonds that are considered polar covalent. Likewise, the Na and Cl atoms in NaCl have an electronegativity difference of 2.1, and the Mn and I atoms in MnI 2 have a difference of 1.0, yet both of these substances form ionic compounds.
The best guide to the covalent or ionic character of a bond is to consider the types of atoms involved and their relative positions in the periodic table. Bonds between two nonmetals are generally covalent; bonding between a metal and a nonmetal is often ionic.
Some compounds contain both covalent and ionic bonds. The atoms in polyatomic ions, such as OH – , and are held together by polar covalent bonds. However, these polyatomic ions form ionic compounds by combining with ions of opposite charge. For example, potassium nitrate, KNO 3 , contains the K + cation and the polyatomic anion. Thus, bonding in potassium nitrate is ionic, resulting from the electrostatic attraction between the ions K + and as well as covalent between the nitrogen and oxygen atoms in
Example \(\PageIndex{1}\): Electronegativity and Bond Polarity
Bond polarities play an important role in determining the structure of proteins. Using the electronegativity values in Figure \(\PageIndex{3}\), arrange the following covalent bonds—all commonly found in amino acids—in order of increasing polarity. Then designate the positive and negative atoms using the symbols δ+ and δ–:
C–H, C–N, C–O, N–H, O–H, S–H
Solution
The polarity of these bonds increases as the absolute value of the electronegativity difference increases. The atom with the δ– designation is the more electronegative of the two. Table \(\PageIndex{1}\) shows these bonds in order of increasing polarity.
| Bond | ΔEN | Polarity |
|---|---|---|
| C–H | 0.4 | |
| S–H | 0.4 | |
| C–N | 0.5 | |
| N–H | 0.9 | |
| C–O | 1.0 | |
| O–H | 1.4 |
Exercise \(\PageIndex{1}\)
Silicones are polymeric compounds containing, among others, the following types of covalent bonds: Si–O, Si–C, C–H, and C–C. Using the electronegativity values in Figure \(\PageIndex{3}\), arrange the bonds in order of increasing polarity and designate the positive and negative atoms using the symbols δ+ and δ–.
- Answer
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Bond Electronegativity Difference Polarity C–C 0.0 nonpolar C–H 0.4 Si–C 0.7 Si–O 1.7


The Acid-Fast Stain
Name: _____________________________________________________
Course Section: ______________________________________________
The following activity will teach students how to complete an Acid-Fast Stain and identify bacteria based on their different cell wall properties. Read the following procedure carefully before starting the activity.
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Materials
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2 microscope slides
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Wax pencil
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Mycobacterium smegmatis
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Staphylococcus epidermidis
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Carbol fuchsin stain
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Methylene blue stain
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Acid alcohol
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DI water
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Bibulous paper
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Staining tray and slide stand
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Inoculation loop
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Hot plate
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Clothespin
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Forceps
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1000 ml beaker
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Timer
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Incinerator
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Microscopes: lens paper, lens cleaner, immersion oil
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Procedure
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Step 1: Prepare A Bacterial Smear
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Get into groups of 2.
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Label the edge of the microscope slide with 1 person's initials and the bacterial species. You may abbreviate the name of the bacteria so it fits on the slide. Try to write small.
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On the bottom of the slide, you may draw a circle in the middle of the slide with a sharpie or wax pencil so you know where to make your smear. This is optional.
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Sterilize your inoculation loop and use it to pick up a small amount of DI water. Place the water on top of the slide, in the middle of the circle if you drew one.
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Sterilize your loop again and pick up a small bacterial colony from the stock plate.
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Gently, mix the bacteria into the water drop on the slide. You want to spread the water and bacteria out to about the size of a dime.
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Let the water and bacteria smear completely air dry. The slide should look dry and "crusty" before moving to Step 2.
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Step 2: Heat Fixation
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Once the bacterial smear is completely dry, use a clothespin to clip the end of your slide.
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Then, place your slide directly on a hot plate for 30 seconds.
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After 30 seconds, place your slide onto the slide stand in your staining tray. Don't touch the slide, it will be hot. Don't place the slide on the cold table, the glass slide could crack.
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Once your slide is completely cool, move on to Step 3.
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Step 3: Acid-Fast Staining
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Primary Staining
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Take your heat-fixed slide to the fume hood (the fumes from carbol fuchsin can be toxic).
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Once the water is boiling, place your slide on the slide rack above the boiling water.
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Cover the smear area of your slide with a square piece of bibulous paper.
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Carefully apply the carbol fuchsin stain to the bibulous paper.
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Steam with the stain on the slide for 5 minutes while continuously applying more stain so the bibulous paper never dries out.
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After 5 minutes remove the paper with forceps and discard it in the trash bin.
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Using a clothespin take your slide back to your lab station and rinse with water.
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Decolorizer, Wash Step
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Hold the slide at a 45° angle and gently rinse with the acid alcohol until the run-off is almost clear.
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Over or under-decolorizing will affect your staining results.
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Wash the decolorizer off the slide with a gentle DI rinse.
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Counterstain
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Add a few drops of methylene blue to the smear.
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Leave the stain for 2 minutes.
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Hold the slide at a 45° angle and rinse with DI water
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Place the slide between 2 sheets of bibulous paper and gently pat dry. Do not wipe the slide to dry it.
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Microscopy
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When the slide is dry, observe your specimen using oil immersion.
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Repeat Steps 1 through 3 for the remaining bacterial species.
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When both slides are done, clean your microscope and place it back in the cabinet.
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Rinse your staining tray in a sink and put all materials back in their labeled locations.
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Results
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Species
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Color
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Acid Fast +/-
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Morphology and Arrangement
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Staphylococcus epidermidis
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Mycobacterium smegmatis
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General Questions
1. In the acid-fast stain procedure, which step is used to differentiate acid-fast positive from acid-fast negative bacteria? Explain your answer.
2. What is the purpose of steam in the acid-fast staining procedure?
3. Describe the structure of the cell walls of acid-fast positive bacteria. How do their cell walls differ from gram-positive and gram-negative cell walls?
4. What is the medical importance of using the acid-fast stain (i.e., what diseases/infections can it help to identify)?


50.6.3: Agglutination Assays
- Compare direct and indirect agglutination
- Identify various uses of hemagglutination in the diagnosis of disease
- Explain how blood types are determined
- Explain the steps used to cross-match blood to be used in a transfusion
In addition to causing precipitation of soluble molecules and flocculation of molecules in suspension, antibodies can also clump together cells or particles (e.g., antigen-coated latex beads) in a process called agglutination ( Figure 18.1.8 ). Agglutination can be used as an indicator of the presence of antibodies against bacteria or red blood cells. Agglutination assays are usually quick and easy to perform on a glass slide or microtiter plate (Figure \(\PageIndex{1}\)). Microtiter plates have an array of wells to hold small volumes of reagents and to observe reactions (e.g., agglutination) either visually or using a specially designed spectrophotometer. The wells come in many different sizes for assays involving different volumes of reagents.
Figure \(\PageIndex{1}\): Microtiter plates are used for conducting numerous reactions simultaneously in an array of wells. (credit: modification of work by “Microrao”/Wikimedia)Agglutination of Bacteria and Viruses
The use of agglutination tests to identify streptococcal bacteria was developed in the 1920s by Rebecca Lancefieldworking with her colleagues A.R. Dochez and Oswald Avery. 1 She used antibodies to identify M protein, a virulence factor on streptococci that is necessary for the bacteria’s ability to cause strep throat. Production of antibodies against M protein is crucial in mounting a protective response against the bacteria.
Lancefield used antisera to show that different strains of the same species of streptococci express different versions of M protein, which explains why children can come down with strep throat repeatedly. Lancefield classified beta-hemolytic streptococci into many groups based on antigenic differences in group-specific polysaccharides located in the bacterial cell wall. The strains are called serovars because they are differentiated using antisera. Identifying the serovars present in a disease outbreak is important because some serovars may cause more severe disease than others.
The method developed by Lancefield is a direct agglutination assay, since the bacterial cells themselves agglutinate. A similar strategy is more commonly used today when identifying serovars of bacteria and viruses; however, to improve visualization of the agglutination, the antibodies may be attached to inert latex beads. This technique is called an indirect agglutination assay (or latex fixation assay), because the agglutination of the beads is a marker for antibody binding to some other antigen (Figure \(\PageIndex{2}\)). Indirect assays can be used to detect the presence of either antibodies or specific antigens.
Figure \(\PageIndex{2}\): Antibodies against six different serovars of Group A strep were attached to latex beads. Each of the six antibody preparations was mixed with bacteria isolated from a patient. The tiny clumps seen in well 4 are indicative of agglutination, which is absent from all other wells. This indicates that the serovar associated with well 4 is present in the patient sample. (credit: modification of work by American Society for Microbiology)To identify antibodies in a patient’s serum, the antigen of interest is attached to latex beads. When mixed with patient serum, the antibodies will bind the antigen, cross-linking the latex beads and causing the beads to agglutinate indirectly; this indicates the presence of the antibody (Figure \(\PageIndex{3}\)). This technique is most often used when looking for IgM antibodies, because their structure provides maximum cross-linking. One widely used example of this assay is a test for rheumatoid factor (RF) to confirm a diagnosis of rheumatoid arthritis. RF is, in fact, the presence of IgM antibodies that bind to the patient’s own IgG. RF will agglutinate IgG-coated latex beads.
In the reverse test, soluble antigens can be detected in a patient’s serum by attaching specific antibodies (commonly mAbs) to the latex beads and mixing this complex with the serum (Figure \(\PageIndex{3}\)).
Agglutination tests are widely used in underdeveloped countries that may lack appropriate facilities for culturing bacteria. For example, the Widal test, used for the diagnosis of typhoid fever, looks for agglutination of Salmonella enterica subspecies typhi in patient sera. The Widal test is rapid, inexpensive, and useful for monitoring the extent of an outbreak; however, it is not as accurate as tests that involve culturing of the bacteria. The Widal test frequently produces false positives in patients with previous infections with other subspecies of Salmonella , as well as false negatives in patients with hyperproteinemia or immune deficiencies.
In addition, agglutination tests are limited by the fact that patients generally do not produce detectable levels of antibody during the first week (or longer) of an infection. A patient is said to have undergone seroconversion when antibody levels reach the threshold for detection. Typically, seroconversion coincides with the onset of signs and symptoms of disease. However, in an HIV infection, for example, it generally takes 3 weeks for seroconversion to take place, and in some instances, it may take much longer.
Similar to techniques for the precipitin ring test and plaque assays, it is routine to prepare serial two-fold dilutions of the patient’s serum and determine the titer of agglutinating antibody present. Since antibody levels change over time in both primary and secondary immune responses, by checking samples over time, changes in antibody titer can be detected. For example, a comparison of the titer during the acute phase of an infection versus the titer from the convalescent phase will distinguish whether an infection is current or has occurred in the past. It is also possible to monitor how well the patient’s immune system is responding to the pathogen.
Figure \(\PageIndex{3}\): (a) Latex beads coated with an antigen will agglutinate when mixed with patient serum if the serum contains IgM antibodies against the antigen. (b) Latex beads coated with antibodies will agglutinate when mixed with patient serum if the serum contains antigens specific to the antibodies.- How is agglutination used to distinguish serovars from each other?
- In a latex bead assay to test for antibodies in a patient's serum, with what are the beads coated?
- What has happened when a patient has undergone seroconversion?
Hemagglutination
Agglutination of red blood cells is called hemagglutination. One common assay that uses hemagglutination is the direct Coombs’ test, also called the direct antihuman globulin test (DAT), which generally looks for nonagglutinating antibodies. The test can also detect complement attached to red blood cells.
The Coombs’ test is often employed when a newborn has jaundice, yellowing of the skin caused by high blood concentrations of bilirubin, a product of the breakdown of hemoglobin in the blood. The Coombs’ test is used to determine whether the child’s red blood cells have been bound by the mother’s antibodies. These antibodies would activate complement, leading to red blood cell lysis and the subsequent jaundice. Other conditions that can cause positive direct Coombs’ tests include hemolytic transfusion reactions, autoimmune hemolytic anemia, infectious mononucleosis (caused by Epstein-Barr virus), syphilis, and Mycoplasma pneumonia. A positive direct Coombs’ test may also be seen in some cancers and as an allergic reaction to some drugs (e.g., penicillin).
The antibodies bound to red blood cells in these conditions are most often IgG, and because of the orientation of the antigen-binding sites on IgG and the comparatively large size of a red blood cell, it is unlikely that any visible agglutination will occur. However, the presence of IgG bound to red blood cells can be detected by adding Coombs’ reagent, an antiserum containing antihuman IgG antibodies (that may be combined with anti-complement) (Figure \(\PageIndex{4}\)). The Coombs’ reagent links the IgG attached to neighboring red blood cells and thus promotes agglutination.
There is also an indirect Coombs’ test known as the indirect antiglobulin test (IAT). This screens an individual for antibodies against red blood cell antigens (other than the A and B antigens) that are unbound in a patient’s serum (Figure \(\PageIndex{4}\)). IAT can be used to screen pregnant women for antibodies that may cause hemolytic disease of the newborn. It can also be used prior to giving blood transfusions. More detail on how the IAT is performed is discussed below.
Figure \(\PageIndex{4}\): The steps in direct and indirect Coombs’ tests are shown in the illustration.Antibodies that bind to red blood cells are not the only cause of hemagglutination. Some viruses also bind to red blood cells, and this binding can cause agglutination when the viruses cross-link the red blood cells. For example, influenza viruses have two different types of viral spikes called neuraminidase (N) and hemagglutinin (H), the latter named for its ability to agglutinate red blood cells (see Viruses ). Thus, we can use red blood cells to detect the presence of influenza virus by direct hemagglutination assays (HA), in which the virus causes visible agglutination of red blood cells. The mumps and rubella viruses can also be detected using HA.
Most frequently, a serial dilution viral agglutination assay is used to measure the titer or estimate the amount of virus produced in cell culture or for vaccine production. A viral titer can be determined using a direct HA by making a serial dilution of the sample containing the virus, starting with a high concentration of sample that is then diluted in a series of wells. The highest dilution producing visible agglutination is the titer. The assay is carried out in a microtiter plate with V- or round-bottomed wells. In the presence of agglutinating viruses, the red blood cells and virus clump together and produce a diffuse mat over the bottom of the well. In the absence of virus, the red blood cells roll or sediment to the bottom of the well and form a dense pellet, which is why flat-bottomed wells cannot be used (Figure \(\PageIndex{5}\)).
A modification of the HA assay can be used to determine the titer of antiviral antibodies. The presence of these antibodies in a patient’s serum or in a lab-produced antiserum will neutralize the virus and block it from agglutinating the red cells, making this a viral hemagglutination inhibition assay (HIA). In this assay, patient serum is mixed with a standardized amount of virus. After a short incubation, a standardized amount of red blood cells is added and hemagglutination is observed. The titer of the patient’s serum is the highest dilution that blocks agglutination (Figure \(\PageIndex{6}\)).
Figure \(\PageIndex{5}\): A viral suspension is mixed with a standardized amount of red blood cells. No agglutination of red blood cells is visible when the virus is absent, and the cells form a compact pellet at the bottom of the well. In the presence of virus, a diffuse pink precipitate forms in the well. (credit bottom: modification of work by American Society for Microbiology) Figure \(\PageIndex{6}\): In this HIA, serum containing antibodies to influenzavirus underwent serial two-fold dilutions in a microtiter plate. Red blood cells were then added to the wells. Agglutination only occurred in those wells where the antibodies were too dilute to neutralize the virus. The highest concentration at which agglutination occurs is the titer of the antibodies in the patient’s serum. In the case of this test, Sample A shows a titer of 128, and Sample C shows a titer of 64. (credit: modification of work by Evan Burkala)- What is the mechanism by which viruses are detected in a hemagglutination assay?
- Which hemagglutination result tells us the titer of virus in a sample?
Much of what we know today about the human immune system has been learned through research conducted using animals—primarily, mammals—as models. Besides research, mammals are also used for the production of most of the antibodies and other immune system components needed for immunodiagnostics. Vaccines, diagnostics, therapies, and translational medicine in general have all been developed through research with animal models.
Consider some of the common uses of laboratory animals for producing immune system components. Guinea pigs are used as a source of complement, and mice are the primary source of cells for making mAbs. These mAbs can be used in research and for therapeutic purposes. Antisera are raised in a variety of species, including horses, sheep, goats, and rabbits. When producing an antiserum, the animal will usually be injected at least twice, and adjuvants may be used to boost the antibody response. The larger animals used for making antisera will have blood harvested repeatedly over long periods of time, with little harm to the animals, but that is not usually the case for rabbits. Although we can obtain a few milliliters of blood from the ear veins of rabbits, we usually need larger volumes, which results in the deaths of the animals.
We also use animals for the study of disease. The only way to grow Treponema pallidum for the study of syphilis is in living animals. Many viruses can be grown in cell culture, but growth in cell culture tells us very little about how the immune system will respond to the virus. When working on a newly discovered disease, we still employ Koch’s postulates, which require causing disease in lab animals using pathogens from pure culture as a crucial step in proving that a particular microorganism is the cause of a disease. Studying the proliferation of bacteria and viruses in animal hosts, and how the host immune system responds, has been central to microbiological research for well over 100 years.
While the practice of using laboratory animals is essential to scientific research and medical diagnostics, many people strongly object to the exploitation of animals for human benefit. This ethical argument is not a new one—indeed, one of Charles Darwin's daughters was an active antivivisectionist (vivisection is the practice of cutting or dissecting a live animal to study it). Most scientists acknowledge that there should be limits on the extent to which animals can be exploited for research purposes. Ethical considerations have led the National Institutes of Health (NIH) to develop strict regulations on the types of research that may be performed. These regulations also include guidelines for the humane treatment of lab animals, setting standards for their housing, care, and euthanization. The NIH document “Guide for the Care and Use of Laboratory Animals” makes it clear that the use of animals in research is a privilege granted by society to researchers.
The NIH guidelines are based on the principle of the three R’s: replace, refine, and reduce. Researchers should strive to replace animal models with nonliving models, replace vertebrates with invertebrates whenever possible, or use computer-models when applicable. They should refine husbandry and experimental procedures to reduce pain and suffering, and use experimental designs and procedures that reduce the number of animals needed to obtain the desired information. To obtain funding, researchers must satisfy NIH reviewers that the research justifies the use of animals and that their use is in accordance with the guidelines.
At the local level, any facility that uses animals and receives federal funding must have an Institutional Animal Care and Use Committee (IACUC) that ensures that the NIH guidelines are being followed. The IACUC must include researchers, administrators, a veterinarian, and at least one person with no ties to the institution, that is, a concerned citizen. This committee also performs inspections of laboratories and protocols. For research involving human subjects, an Institutional Review Board (IRB) ensures that proper guidelines are followed.
Blood Typing and Cross-Matching
In addition to antibodies against bacteria and viruses to which they have previously been exposed, most individuals also carry antibodies against blood types other than their own. There are presently 33 immunologically important blood-type systems, many of which are restricted within various ethnic groups or rarely result in the production of antibodies. The most important and perhaps best known are the ABO and Rh blood groups (see Figure 19.1.3 ).
When units of blood are being considered for transfusion, pretransfusion blood testing must be performed. For the blood unit, commercially prepared antibodies against the A, B, and Rh antigens are mixed with red blood cells from the units to initially confirm that the blood type on the unit is accurate. Once a unit of blood has been requested for transfusion, it is vitally important to make sure the donor (unit of blood) and recipient (patient) are compatible for these crucial antigens. In addition to confirming the blood type of the unit, the patient’s blood type is also confirmed using the same commercially prepared antibodies to A, B, and Rh. For example, as shown in Figure \(\PageIndex{7}\), if the donor blood is A-positive, it will agglutinate with the anti-A antiserum and with the anti-Rh antiserum. If no agglutination is observed with any of the sera, then the blood type would be O-negative.
Following determination of the blood type, immediately prior to releasing the blood for transfusion, a cross-match is performed in which a small aliquot of the donor red blood cells are mixed with serum from the patient awaiting transfusion. If the patient does have antibodies against the donor red blood cells, hemagglutination will occur. To confirm any negative test results and check for sensitized red blood cells, Coombs’ reagent may be added to the mix to facilitate visualization of the antibody-red blood cell interaction.
Under some circumstances, a minor cross-match may be performed as well. In this assay, a small aliquot of donor serum is mixed with patient red blood cells. This allows the detection of agglutinizing antibodies in the donor serum. This test is rarely necessary because transfusions generally use packed red blood cells with most of the plasma removed by centrifugation.
Red blood cells have many other antigens in addition to ABO and Rh. While most people are unlikely to have antibodies against these antigens, women who have had multiple pregnancies or patients who have had multiple transfusions may have them because of repeated exposure. For this reason, an antibody screen test is used to determine if such antibodies are present. Patient serum is checked against commercially prepared, pooled, type O red blood cells that express these antigens. If agglutination occurs, the antigen to which the patient is responding must be identified and determined not to be present in the donor unit.
Figure \(\PageIndex{7}\): This sample of a commercially produced “bedside” card enables quick typing of both a recipient’s and donor’s blood before transfusion. The card contains three reaction sites or wells. One is coated with an anti-A antibody, one with an anti-B antibody, and one with an anti-Rh antibody. Agglutination of red blood cells in a given site indicates a positive identification of the blood antigens: in this case, A and Rh antigens for blood type A-positive.- If a patient's blood agglutinates with anti-B serum, what is the patient’s blood type?
- What is a cross-match assay, and why is it performed?
Table \(\PageIndex{1}\) summarizes the various kinds of agglutination assays discussed in this section.
| Type of Assay | Mechanism | Example |
|---|---|---|
| Agglutination | Direct: Antibody is used to clump bacterial cells or other large structures | Serotyping bacteria |
| Indirect: Latex beads are coupled with antigen or antibody to look for antibody or antigen, respectively, in patient serum | Confirming the presence of rheumatoid factor (IgM-binding Ig) in patient serum | |
| Hemagglutination | Direct: Some bacteria and viruses cross-link red blood cells and clump them together | Diagnosing influenza, mumps, and measles |
| Direct Coombs’ test (DAT): Detects nonagglutinating antibodies or complement proteins on red blood cells in vivo | Checking for maternal antibodies binding to neonatal red blood cells | |
| Indirect Coombs’ test (IAT): Screens an individual for antibodies against red blood cell antigens (other than the A and B antigens) that are unbound in a patient’s serum in vitro | Performing pretransfusion blood testing | |
| Viral hemagglutination inhibition: Uses antibodies from a patient to inhibit viral agglutination | Diagnosing various viral diseases by the presence of patient antibodies against the virus | |
| Blood typing and cross-matching: Detects ABO, Rh, and minor antigens in the blood | Matches donor blood to recipient immune requirements |
Key Concepts and Summary
- Antibodies can agglutinate cells or large particles into a visible matrix. Agglutination tests are often done on cards or in microtiter plates that allow multiple reactions to take place side by side using small volumes of reagents.
- Using antisera against certain proteins allows identification of serovars within species of bacteria.
- Detecting antibodies against a pathogen can be a powerful tool for diagnosing disease, but there is a period of time before patients go through seroconversion and the level of antibodies becomes detectable.
- Agglutination of latex beads in indirect agglutination assays can be used to detect the presence of specific antigens or specific antibodies in patient serum.
- The presence of some antibacterial and antiviral antibodies can be confirmed by the use of the direct Coombs’ test , which uses Coombs’ reagent to cross-link antibodies bound to red blood cells and facilitate hemagglutination .
- Some viruses and bacteria will bind and agglutinate red blood cells; this interaction is the basis of the direct hemagglutination assay , most often used to determine the titer of virus in solution.
- Neutralization assays quantify the level of virus-specific antibody by measuring the decrease in hemagglutination observed after mixing patient serum with a standardized amount of virus.
- Hemagglutination assays are also used to screen and cross-match donor and recipient blood to ensure that the transfusion recipient does not have antibodies to antigens in the donated blood.
Footnotes
- 1 Lancefield, Rebecca C., “The Antigenic Complex of Streptococcus haemoliticus . I. Demonstration of a Type-Specific Substance in Extracts of Streptococcus haemolyticus ,” The Journal of Experimental Medicine 47, no. 1 (1928): 91-103.


6.8: Dilworth's Theorem for Interval Orders
As remarked previously, we do not yet have an efficient process for determining the width of a poset and a minimum partition into chains. For interval orders, there is indeed a simple way to find both. The explanation is just to establish a connection with coloring of interval graphs as discussed in Chapter 5.
Let \(\textbf{P}=(X,P)\) be an interval order and let \(\{[a_x,b_x]:x \in X\}\) be intervals of the real line so that \(x<y\) in \(\textbf{P}\) if and only \(b_x<a_y\). Then let \(\textbf{G}\) be the interval graph determined by this family of intervals. Note that if \(x\) and \(y\) are distinct elements of \(X\), then \(x\) and \(y\) are incomparable in \(\textbf{P}\) if and only if \(xy\) is an edge in \(\textbf{G}\). In other words, \(\textbf{G}\) is just the incomparability graph of \(\textbf{P}\).
Recall from Chapter 5 that interval graphs are perfect, i.e., \(\chi(\textbf{G})=ω(\textbf{G})\) for every interval graph \(\textbf{G}\). Furthermore, you can find an optimal coloring of an interval graph by applying first fit to the vertices in a linear order that respects left end points. Such a coloring concurrently determines a partition of \(\textbf{P}\) into chains.
In fact, if you want to skip the part about interval representations, take any linear ordering of the elements as \(x_1, x_2,…,x_n\) so that \(i<j\) whenever \(D(x)\) is a proper subset of \(D(y)\). Then apply First Fit with respect to chains. For example, using the 10 point interval order illustrated in Figure 6.31 , here is such a labeling:
\(x_1=g\) \(x_2=f\) \(x_3=c\) \(x_4=d\) \(x_5=h\)
\(x_6=a\) \(x_7=j\) \(x_8=b\) \(x_9=i\) \(x_{10}=e\)
Now apply the First Fit algorithm to the points of \(\textbf{P}\), in this order, to assign them to chains \(C_1, C_2,….\) In other words, assign \(x_1\) to chain \(C_1\). Thereafter if you have assigned points \(x_1, x_2,…,x_i\) to chains, then assign \(x_{i+1}\) to chain \(C_j\) where \(j\) is the least positive integer for which \(x_{i+1}\) is comparable to \(x_k\) whenever \(1 \leq k \leq i\) and \(x_k\) has already been assigned to \(C_j\). For example, this rule results in the following chains for the interval order \(\textbf{P}\) shown in Figure 6.31 .
\(C_1 = \{g,h,b\}\)
\(C_2 = \{f,a,e\}\)
\(C_3 = \{c,d\}\)
\(C_4 = \{j\}\)
\(C_5 = \{i\}\)
In this case, it is easy to see that the chain partition is optimal since the width of \(\textbf{P}\) is 5 and \(A=\{a,b,d,i,j\}\) is a 5-element antichain.
However, you should be very careful in applying First Fit to find optimal chain partitions of posets—just as one must be leary of using First Fit to find optimal colorings of graphs.
The poset on the left side of Figure 6.33 is a height 2 poset on 10 points, and if the poset is partitioned into antichains by applying First Fit and considering the points in the order of their labels, then 5 antichains will be used. Do you see how to extend this poset to force First Fit to use arbitrarily many antichains, while keeping the height of the poset at 2?
On the right side, we show a poset of width 2. Now if this poset is partitioned into chains by applying First Fit and considering the points in the order of their labels, then 4 chains will be used. Do you see how to extend this poset to force First Fit to use arbitrarily many chains while keeping the width of the poset at 2?
Do you get a feeling for why the second problem is a bit harder than the first?
In general, there is always some linear order on the ground set of a poset for which First Fit will find an optimal partition into antichains. Also, there is a linear order (in general different from the first) on the ground set for which First Fit will find an optimal partition into chains. However, there is no advantage in searching for such orders, as the algorithms we develop for finding optimal antichain and chain partitions work quite well.


30.6B: Development of the Peripheral Nervous System
The peripheral nervous system develops from two strips of tissue called the neural crest, running lengthwise above the neural tube.
- Describe peripheral nervous system development
Key Points
- The first sign of the nervous system is the appearance of a thin strip of cells along the center of the back, called the neural plate. The inner portion of the neural plate is destined to become the central nervous system (CNS), while the outer portion will become the peripheral nervous system (PNS).
- Neurulation (neural development) progresses with the formation of the neural groove which closes to form the neural tube and neural crest.
- Neural crest cells from the roof plate of the neural tube migrate through the periphery where they differentiate into varied cell types, including pigment cells and the cells of the PNS.
Key Terms
- neurulation : The process by which the beginnings of the vertebrate nervous system is formed in embryos.
- neural plate : A thick, flat bundle of ectoderm formed in vertebrate embryos after induction by the notochord.
- neural crest : A strip of ectodermal material in the early vertebrate embryo inserted between the prospective neural plate and the epidermis.
In vertebrates, the first sign of the nervous system is the appearance of a thin strip of cells along the center of the back, called the neural plate. The inner portion of the neural plate (along the midline) is destined to become the central nervous system (CNS), the outer portion the peripheral nervous system (PNS). As development proceeds, a fold called the neural groove appears along the midline. This fold deepens and then closes up at the top. At this point the future CNS appears as a cylindrical structure called the neural tube, whereas the future PNS appears as two strips of tissue called the neural crest, running lengthwise above the neural tube. The sequence of stages from neural plate to neural tube and neural crest is known as neurulation.
Formation of the Fetal Nervous System : The neural tube will give rise to the central nervous system, while the neural crest will give rise to the peripheral nervous system. This diagram shows the epidermis, neural fold, and convergence.
Neural crest cells are a transient, multipotent, migratory cell population unique to vertebrates that gives rise to a diverse cell lineage including melanocytes, craniofacial cartilage and bone, smooth muscle, peripheral and enteric neurons and glia. After gastrulation, neural crest cells are specified at the border of the neural plate and the non-neural ectoderm. During neurulation, the borders of the neural plate, also known as the neural folds, converge at the dorsal midline to form the neural tube. Subsequently, neural crest cells from the roof plate of the neural tube undergo an epithelial to mesenchymal transition, delaminating from the neuroepithelium and migrating through the periphery where they differentiate into varied cell types, including pigment cells and the cells of the PNS. The emergence of the neural crest was important in vertebrate evolution because many of its structural derivatives are defining features of the vertebrate clade.


6.5: The Atkins Diet and Gluconeogenesis
You may know that the Atkins Diet is an ultra-low carb diet. It is one of several low-carb ketogenic diets . The glucocorticoid hormones released on a low carb diet trick the body into a constant gluconeogenic state. While the liver can produce enough glucose for brain and heart cells, the rest of the cells in our bodies switch to burning fats, hence the weight loss. Discredited some years ago, the Atkins Diet (and similar ones e.g., South Beach) is now back in favor. Some folks on these diets restrict their intake of carbohydrates so much that they can develop “acetone breath”! Nevertheless, low carb diets are important in the control of diabetes. In older folks, type 2 (adult-onset) diabetics can control their disease with a low carb diet and a drug called metformin , which blocks gluconeogenesis and therefore prevents glucose synthesis from gluconeogenic substrates, at the same time stimulating cellular receptors to take up available glucose. For more details on the mechanism of metformin action, check out Hundal RS et al. [(2000) Mechanism by Which Metformin Reduces Glucose Production in Type 2 Diabetes. Diabetes 49 (12): 2063–9]. Given the prevalence of obesity and type 2 diabetes in the U.S., it’s likely that someone you know is taking metformin or other similar medication!


21.12: Checklist for Obtaining a Urine Specimen from a Foley Catheter
Use the checklist below to review the steps for completion of “Obtaining a Urine Specimen from a Foley Catheter.”
Steps
Disclaimer: Always review and follow agency policy regarding this specific skill.
- Gather supplies: peri-care supplies, nonsterile gloves, 30 – 60 mL Luer-lock syringe for sterile specimen, alcohol wipes/scrub hubs, two preprinted patient labels, clear biohazard bag for lab sample, urinary graduated cylinder, and 10-mL syringe.
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Perform safety steps:
- Perform hand hygiene.
- Check the room for transmission-based precautions.
- Introduce yourself, your role, the purpose of your visit, and an estimate of the time it will take.
- Confirm patient ID using two patient identifiers (e.g., name and date of birth).
- Explain the process to the patient.
- Be organized and systematic.
- Use appropriate listening and questioning skills.
- Listen and attend to patient cues.
- Ensure the patient’s privacy and dignity.
- Assess ABCs.
- Verify the order and assemble the supplies on a protective drape on the table: gloves, Luer-lock syringe, alcohol swabs, sterile container, two preprinted patient labels, and clear lab specimen biohazard bag for transport to lab.
- Perform hand hygiene and put on nonsterile gloves.
- Check for urine in the tubing and position the tubing on the bed.
- If additional urine is needed, clamp the tubing below the port for 10-15 minutes or until urine appears.
- Clean the sample port of the catheter with an alcohol swab.
- Attach the Luer-lock syringe to the sample port of the catheter and withdraw 10-30 mL of urine; remove the syringe and unclamp the tubing.
- Open the lid of the sterile container, inverting the lid on the drape and maintaining sterility. Transfer the urine to the sterile container, preventing touching the syringe to the container; place the syringe on the drape; close the lid tightly; clean the outside of the container with germicidal wipes.
- Remove gloves and perform hand hygiene.
- Add information to the preprinted label: date, time collected, and your initials. Apply gloves. Place the label on the specimen container and put the container inside the biohazard bag. Remove gloves and wash your hands. Place the second label outside of the bag. Transport to the lab immediately.
- Assist the patient to a comfortable position, ask if they have any questions, and thank them for their time.
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Ensure safety measures when leaving the room:
- CALL LIGHT: Within reach
- BED: Low and locked (in lowest position and brakes on)
- SIDE RAILS: Secured
- TABLE: Within reach
- ROOM: Risk-free for falls (scan room and clear any obstacles)
- Perform hand hygiene.
- Document the procedure and related assessment findings. Report any concerns according to agency policy.