Early in his career, Kai Simons pursued research in the field of medical biochemistry. Both his master's thesis and postgraduate research focused on vitamin B12 absorption. After returning from his post-doc scholarship he continued research on vitamin B12 as well as on blood plasma proteins, but soon started investigating Semliki Forest virus, focusing on its membrane and its lipid composition and their role in the virus budding and its transport, as the model for lipid and protein secretion. During this period, Simons also investigated the application of detergents in biochemistry with a special attention to their role in biological membrane research.
The virus lifecycle and how it uses components of vesicular pathways while shuttling to the cell surface, turned Simons' attention toward vesicular transport pathways and cell polarization. Applying epithelial model cells – MDCK (Madin-Darby canine kidney), he investigated lipid transport, protein sorting and their role in polarizing cells.
In these studies, he described the role of the trans-Golgi network (TGN) in protein and lipid sorting according to their destination. In his works from 1988, together with Gerit van Meer, Simons proposed the existence of lipid microdomains in cell membranes for the first time. Such microdomains differ in their composition from the surrounding membrane and have special functions. Simons coined the term lipid rafts to describe these microdomains. This concept was developed over the years to be presented more fully in 1997 in Nature journal by Simons and Ikonen. This paper became one of the most frequently cited works in the field of membrane research. Other Simons' paper, on role of lipid rafts in the signal transduction is second highest cited work and Simons is fourth highest cited scientist in the field of signal transduction. Kai Simons was also recognized by ISI Web of Knowledge, as one of the most cited scientist ever.
In subsequent years, Simons continued to work on the role of lipid rafts, and more generally lipids, in cell polarization and protein sorting. He was interested also in the role of lipids and protein sorting in neurodegenerative diseases, especially in Alzheimer's disease.
His scientific record includes more than 350 scientific articles, mostly in the field of biochemistry, molecular organization of the cell, and biochemistry and physiology of a cell membrane.
Considering his work from years 1996–2007 tracked until May 2009, Simons was 12. in the list of the most frequently cited scientists in the field cell biology with 90 articles and 16,299 citations.