With this review, we aim to introduce scientists working in the field of Quaternary paleolimnology to the tools that compound-specific isotope analysis (CSIA) provides for the gain of information on biogeochemical conditions in ancient environments. We provide information on fundamental principles and applications of novel and established CSIA applications based on the carbon, hydrogen, nitrogen, oxygen and sulfur isotopic composition of biomarkers. While biosynthesis, sources and associated isotope fractionation patterns of compounds such as n-alkanes are relatively well-constrained, new applications emerge from the increasing use of functionalized alkyl lipids, steroids, hopanoids, isoprenoids, GDGTs, pigments or cellulose. Biosynthesis and fractionation are not always fully understood. However, although analytical challenges remain, the future potential of deeper insights into ecosystem dynamics from the study of these compounds is also emerging.