Immunoaffinity chromatography of lymphocyte subpopulations using tert-amine derived matrices with adsorbed antibodies.
New polymeric solid-phase matrices for cell affinity chromatography were prepared and their advantageous characteristics compared with conventional matrices were highlighted. These new matrices are derivatives of poly(2-hydroxyethyl methacrylate) (PHEMA) containing a slight quantity of amino compounds as a co-monomer. They were applied to immunoaffinity selection between IgG+ and IgG- lymphocytes of the rat mesenteric lymph node. Simple physical adsorption was sufficient for anti-rat IgG antibodies to be immobilized on these matrices, allowing us to omit the laborious procedure of covalent-linking of antibodies on a matrix. As these matrices themselves showed extremely low non-specific adsorption of lymphocytes, a very dilute solution of antibody (0.02-0.08 mg/ml) was enough for column conditioning. This separation method gave IgG- lymphocytes of more than 90% purity and almost 95% yield within as short a time as 7 min. Further, IgG+ lymphocytes were obtained in good yield (80-90% of loaded number) by recovering the adsorbing cell fraction from the column by gentle pipetting of the matrix.
Kataoka
K
K
Institute of Biomedical Engineering, Tokyo Women's Medical College, Japan.
Sakurai
Y
Y
Hanai
T
T
Maruyama
A
A
Tsuruta
T
T
D000588:Amines; D000818:Animals; D000906:Antibodies; D002469:Cell Separation / Q000379:methods; D002846:Chromatography, Affinity / Q000379:methods; D007074:Immunoglobulin G; D008198:Lymph Nodes / Q000276:immunology; D008214:Lymphocytes / Q000145:classification* / Q000166:cytology / Q000276:immunology; D008297:Male; D011102:Polyhydroxyethyl Methacrylate; D051381:Rats; D011919:Rats, Inbred Strains
D000588:Amines; D000906:Antibodies; D007074:Immunoglobulin G; D011102:Polyhydroxyethyl Methacrylate