Fractionation of human lymphocyte subpopulations on immunoglobulin coated Petri dishes. 1981

S Fong, and C D Tsoukas, and J L Pasquali, and R I Fox, and J E Rose, and D Raiklen, and D A Carson, and J H Vaughan

This report describes a simple solid-phase technique for the positive selection of lymphocytes labeled with fluoresceinated antibodies. B lymphocytes were labeled with fluoresceinated anti-human Ig or monoclonal anti-human Ia (L243), and then were bound to plastic culture dishes coated with affinity purified goat anti-fluorescein specific antibody. Bound cells were eluted at 37 degrees C with 1 mM fluorescein-L-lysine phosphate-buffered saline. Functionally the eluted Ig positive cells responded to pokeweed mitogen (PWM) by in vitro secretion of IgM, as measured by radioimmunoassay of culture supernatants. The secretion of IgM was dependent on the addition of T lymphocytes. Moreover, the isolated B cells were functionally receptive to 'help' and 'suppression' by T cells with and without Fc receptors for IgG respectively. T cell subsets were fractionated on plastic culture dishes coated with heat aggregated rabbit or human IgG. the non-bound cells (enriched T(gamma-)) provided collaborative 'help' in the PWM induced IgM secretion response by human B lymphocytes. The bound cells (enriched T(gamma+)) eluted with 0.01 M EDTA in phosphate-buffered saline, suppressed IgM secretion. This method can be adapted to fractionate subsets of lymphocytes for which a fluoresceinated antibody is available. For routine functional studies, the isolation of cell types with conventional or monoclonal antibodies does not require the use of a fluorescence activated cell sorter, but only an antifluorescein labeled Petri dish. In conclusion, a rapid solid-phase technique enables us to prepare enriched populations of functionally active lymphocytes.

UI MeSH Term Description Entries
D007136 Immunoglobulins Multi-subunit proteins which function in IMMUNITY. They are produced by B LYMPHOCYTES from the IMMUNOGLOBULIN GENES. They are comprised of two heavy (IMMUNOGLOBULIN HEAVY CHAINS) and two light chains (IMMUNOGLOBULIN LIGHT CHAINS) with additional ancillary polypeptide chains depending on their isoforms. The variety of isoforms include monomeric or polymeric forms, and transmembrane forms (B-CELL ANTIGEN RECEPTORS) or secreted forms (ANTIBODIES). They are divided by the amino acid sequence of their heavy chains into five classes (IMMUNOGLOBULIN A; IMMUNOGLOBULIN D; IMMUNOGLOBULIN E; IMMUNOGLOBULIN G; IMMUNOGLOBULIN M) and various subclasses. Globulins, Immune,Immune Globulin,Immune Globulins,Immunoglobulin,Globulin, Immune
D007158 Immunologic Techniques Techniques used to demonstrate or measure an immune response, and to identify or measure antigens using antibodies. Antibody Dissociation,Immunologic Technic,Immunologic Technics,Immunologic Technique,Immunological Technics,Immunological Techniques,Technic, Immunologic,Technics, Immunologic,Technique, Immunologic,Techniques, Immunologic,Antibody Dissociations,Dissociation, Antibody,Dissociations, Antibody,Immunological Technic,Immunological Technique,Technic, Immunological,Technics, Immunological,Technique, Immunological,Techniques, Immunological
D008214 Lymphocytes White blood cells formed in the body's lymphoid tissue. The nucleus is round or ovoid with coarse, irregularly clumped chromatin while the cytoplasm is typically pale blue with azurophilic (if any) granules. Most lymphocytes can be classified as either T or B (with subpopulations of each), or NATURAL KILLER CELLS. Lymphoid Cells,Cell, Lymphoid,Cells, Lymphoid,Lymphocyte,Lymphoid Cell
D011817 Rabbits A burrowing plant-eating mammal with hind limbs that are longer than its fore limbs. It belongs to the family Leporidae of the order Lagomorpha, and in contrast to hares, possesses 22 instead of 24 pairs of chromosomes. Belgian Hare,New Zealand Rabbit,New Zealand Rabbits,New Zealand White Rabbit,Rabbit,Rabbit, Domestic,Chinchilla Rabbits,NZW Rabbits,New Zealand White Rabbits,Oryctolagus cuniculus,Chinchilla Rabbit,Domestic Rabbit,Domestic Rabbits,Hare, Belgian,NZW Rabbit,Rabbit, Chinchilla,Rabbit, NZW,Rabbit, New Zealand,Rabbits, Chinchilla,Rabbits, Domestic,Rabbits, NZW,Rabbits, New Zealand,Zealand Rabbit, New,Zealand Rabbits, New,cuniculus, Oryctolagus
D002448 Cell Adhesion Adherence of cells to surfaces or to other cells. Adhesion, Cell,Adhesions, Cell,Cell Adhesions
D002469 Cell Separation Techniques for separating distinct populations of cells. Cell Isolation,Cell Segregation,Isolation, Cell,Cell Isolations,Cell Segregations,Cell Separations,Isolations, Cell,Segregation, Cell,Segregations, Cell,Separation, Cell,Separations, Cell
D005452 Fluoresceins A family of spiro(isobenzofuran-1(3H),9'-(9H)xanthen)-3-one derivatives. These are used as dyes, as indicators for various metals, and as fluorescent labels in immunoassays. Tetraiodofluorescein
D006358 Hot Temperature Presence of warmth or heat or a temperature notably higher than an accustomed norm. Heat,Hot Temperatures,Temperature, Hot,Temperatures, Hot
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia

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