Abstract
A method is presented for the rapid determination of the fluorescein content, protein content, and fluorescein-to-protein ratio for immune globulin conjugates with fluorescein isothiocyanate as the fluor. This method is based on the absorbance of the fluorescent antibody at those wavelengths primarily associated with the fluorescein and γ-globulin fractions, and permits these materials to be determined by a single nondestructive analytical procedure. A small sample of the fluorescent antibody, in many cases 0.1 ml or less, is adequate for the above determinations. A nomograph is presented which allows simultaneous determination of the materials from the observed absorbance at the two wavelengths. The method is sufficiently accurate for most applications of the fluorescent-antibody techniques. Although this procedure has been developed primarily for fluorescent-antibody conjugates prepared from rabbit γ-globulin, it can be used directly for antibodies prepared from other animals provided the γ-globulin is relatively free from albumin.
Full text
PDF
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- KABAT E. A., MCDUFFIE F. C. A comparative study of methods used for analysis of specific precipitates in quantitative immunochemistry. J Immunol. 1956 Sep;77(3):193–197. [PubMed] [Google Scholar]
- LEWIS V. J., JONES W. L., BROOKS J. B., CHERRY W. B. TECHNICAL CONSIDERATIONS IN THE PREPARATION OF FLUORESCENT-ANTIBODY CONJUGATES. Appl Microbiol. 1964 Jul;12:343–348. doi: 10.1128/am.12.4.343-348.1964. [DOI] [PMC free article] [PubMed] [Google Scholar]
- MCKINNEY R. M., SPILLANE J. T., PEARCE G. W. DETERMINATION OF PURITY OF FLUORESCEIN ISOTHIOCYANATES. Anal Biochem. 1964 Jan;7:74–86. doi: 10.1016/0003-2697(64)90121-6. [DOI] [PubMed] [Google Scholar]
- MCKINNEY R. M., SPILLANE J. T., PEARCE G. W. FACTORS AFFECTING THE RATE OF REACTION OF FLUORESCEIN ISOTHIOCYANATE WITH SERUM PROTEINS. J Immunol. 1964 Aug;93:232–242. [PubMed] [Google Scholar]