Xp. Li et al., O-PHTHALDEHYDE FLUORESCENCE MICROASSAY FOR THE DETERMINATION OF ANTIBODY CONCENTRATION, Journal of immunological methods, 172(2), 1994, pp. 141-145
o-Phthaldehyde (OPA) reagent reacts with primary amines in aqueous sol
ution at room temperature. When the reaction occurs in the presence of
2-mercaptoethanol, a bright blue fluorescence is produced. We investi
gated the application of the OPA reagent in the routine determination
of monoclonal antibody concentration. The OPA reagent produced maximal
fluorescence within 30 min and was stable for 2 h. Buffers containing
primary amines gave high background fluorescence; however, the OPA re
agent gave reliable results with the most commonly used antibody solut
ions. The OPA reagent provided a rapid and simple measure of protein c
oncentration from 1 mu g to 1 mg and was readily adapted for use with
a 96-well fluorescence reader.