Biotinylation of human interleukin-2 for flow cytometry analysis of interleukin-2 receptors.
We designed a method to analyze receptors for interleukin-2 (IL-2R) using biotinylated IL-2 (b-IL-2). To optimize the condition of biotinylation of IL-2 for flow cytometry, the degree of biotinylation was controlled by monitoring the relative biotin contents in b-IL-2 with a newly developed ELISA. The b-IL-2 prepared by incubating 150 micrograms IL-2 in 150-300 micrograms/ml N-hydroxysuccinimidyl biotin retained biological activity and was appropriate for flow cytometry analysis. Positive fluorescence appeared in the IL-2R-bearing cell lines but not in those without IL-2R. This binding was inhibited by preincubation of the cells with unlabelled IL-2. The b-IL-2 bound to both low and high affinity IL-2Rs, but the binding to the latter was more intense. The advantage of this method is that expression of IL-2Rs of these two categories of affinity can be separately monitored.
Taki
S
S
Central Research Laboratory, Ajinomoto Co., Inc., Kawasaki, Japan.
Shimamura
T
T
Abe
M
M
Shirai
T
T
Takahara
Y
Y
D000915:Antibody Affinity; D000937:Antigen-Antibody Reactions; D001667:Binding, Competitive; D001710:Biotin; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005434:Flow Cytometry; D006801:Humans; D007158:Immunologic Techniques; D007376:Interleukin-2 / Q000276:immunology* / Q000378:metabolism; D007700:Kinetics; D008214:Lymphocytes / Q000032:analysis; D011485:Protein Binding; D015375:Receptors, Interleukin-2 / Q000032:analysis* / Q000276:immunology / Q000378:metabolism
D007376:Interleukin-2; D015375:Receptors, Interleukin-2; D001710:Biotin