The previously unknown difluoromethyl-and the previously neglected trifluoromethyl-aliphatic diazirines were synthesized and characterized. Model photolabeling experiments and biological studies revealed that the stable trifluoromethyl-aliphatic diazirines could indeed be safely used as photoaffinity labels, while the labile difluoromethylated analogues -could not. Introduction and Aim. Aliphatic diazirines (type A, Figure ) were discovered first in the 1960s by Abendroth with Henrich; Paulsen; and Schmitz with Ohme. Their thermal decomposition was originally studied as an approach to carbenes. Photochemical irradiation of diazirines also leads to the elimination of nitrogen and the formation of carbenes. The latter can insert at the C-H and N-H, C-H, S-H bonds to cross-link with proximal species. Therefore, aliphatic diazirines have become common labels for photoaffinity studies. It is worth noting that aliphatic diazirines tend to undergo the undesired 1,2-hydrogen rearrangement into alkenes. In 1973, Knowles studied the utility of aromatic diazirines for photolabeling. Under irradiation, however, they extensively isomerized into the more stable diazo compounds that caused the nonspecific labeling. In 1980, Brunner demonstrated that the trifluoromethyl-substituted aromatic diazirines (type B, Figure ) were more useful as photoaffinity labels, because the undesired isomerization products -CF 3 -diazo compounds CF 3 C(N 2 )Ar,were inert and did not cause the non-selective labeling. Since that time, chemists have been commonly using trifluoromethylsubstituted aromatic diazirines as labels to replace aromatic fragments in bioactive compounds for photoaffinity studies. Both types of diazirines -dialkyl and trifluoromethyl-aryl (types A and B, Figure ) -are popular today. In 1984, Meese showed that in strict contrast to diazomethane or alkyl diazomethanes (CH 2 N 2 , MeCHN 2 ), trifluorodiazoethane (CF 3 CHN 2 ) did not react with carboxylic acids. Therefore, we wondered if aliphatic fluoroalkyl-substituted diazirines (type C, Figure ) could be used as photoaffinity labels. The side photoisomerization products -fluoroalkyl diazo compounds,should not cause the undesired site-selective labeling, due to lower activity. Unexpectedly, chemists almost did not use aliphatic trifluoromethyl diazirines before. Moreover, these compounds were mostly unknown from the synthetic standpoint. We could find only three single molecules in the literature with a linear aliphatic CF 3 -diazirine motif, while cyclic molecules were not reported. Aliphatic difluoromethyl-substituted diazirines were completely unknown. Presumably, the lack of synthetic efforts to fluoroalkyl aliphatic diazirines led to the inability of biochemists to systematically study them, and use them as photoaffinity labels. No one made them, and no one could use them.