Preparation of Fluorescein Isothiocyanate-Labeled γ-Globulin by Dialysis, Gel Filtration, and Ion-Exchange Chromatography in Combination

Abstract

DEDMON, ROBERT E. (Presbyterian-St. Luke's Hospital, Chicago, Ill.), ALBERT W. HOLMES, AND FRIEDRICH DEINHARDT. Preparation of fluorescein isothiocyanate-labeled 7-globulin by dialysis, gel filtration, and ion-exchange chromatography in combination. J. Bacteriol. 89:734-739. 1965.-Antiviral immune 7y-globulins isolated from rabbit and guinea pig sera were labeled through dialysis membranes with fluorescein isothiocyanate and purified in several ways to eliminate nonspecific staining. Gel filtration of the conjugate with Sephadex G-25 coarse beads followed by column fractionation with diethylaminoethyl-Sephadex yielded consistently highly specific staining materials. Fluorescein-protein ratios varied between 1.0 and 4.0. This technique has proved to be simple and reliable, and is less time-consuming than previous techniques.

Conjugates of Immunoglobulin G with Different Fluorochromes. I. Characterization by Anionic‐exchange Chromatography

“…The latter procedure is preferred since it is convenient, and because a protein loss of about 10 per cent as well as a major increase of sample volume can be avoided. Subsequent chromatography may be carried out on columns of DEAH-Sephadex A-50 (12) or DEAE cellulose. Only small differences were noted between the three types of cellulose used (Table II, Nos.…”

A Method for Staining Intracellular Antigens in Thin Sections With Ferritin-Labeled Antibody

“…Purified Microccus sodonensis cell wall preparations (less than 0.01% phosphate) and rabbit antisera to the cell walls were obtained from Dr. J. N. Campbell, Department of Microbiology, University of Alberta. In all eases the antibody was separated in the T-globulin fraction from the sera by passage through a diethylaminoethyl (DEAE)-sephadex column (Dedmon et al, 1965). Ferrltin (6 X crystallized, Cd-free; Polysciences, Inc., Rydal, Pa.) was conjugated to the T-globulin by using toluene-2,4 diisocyanate (Singer and Schick, 1961).…”

Latex agglutination test for detection of Clostridium difficile toxin in stool samples

“…Latex agglutination test. Antitoxin obtained from T. D. Wilkins was passed through a column of DEAE-Sephadex A50 (Pharmacia Fine Chemicals, Piscataway, N.J.) by the method of Dedmon et al (5). Samples (3 ml) of antitoxin were applied, and the column was eluted with 0.02 M phosphate buffer (pH 7.6).…”