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M Mitard

Publications and source records attributed to M Mitard.

5 recordsLinked to original sources
PubMed1976

Immunoenzymatic methods for the study of surface bound immunoglobulins in mice.

Methods are described for the study of lymphocyte surface bound immunoglobulins (Ig) for light microscopy experimentation. a) Live peripheral lymph node cells (LNC) or spleen cells were reacted in suspension under capping conditions with rabbit anti-mouse Ig antibodies. After washing the cells were cytocentrifuged, fixed and relabeled with either peroxidase-(PO) of fluorescein-(Fl) conjugated anti-rabbit IgG antibodies (Method SC). The staining was heterogeneous and three main categories of lymphocytes were distinguished on the basis of a characteristic surface label distribution (1) all label within distinct caps (caps), (2) all label evenly distributed over the cell surface (rings), and (3) an intermediate pattern where only part of the label was concentrated over one pole (polarized). b) LNC or spleen cells were treated for the simultaneous detection of lymphocyte surface Ig and cytoplasmic Ig of plasma cells. For this, live cells were cytocentrifuged, and after fixation were labeled with PO-conjugated antibodies. Lymphocytes stained in a continuous ring and were easily distinguished on morphological grounds from the intense cytoplasmic stain of plasma cells (Method D). Positive lymphocyte counts done by this method compared favorably with those obtained by Method SC. Comparison of Fl- and PO-conjugated reagents were done by either Method SC or D, by scoring total positive cells. Although either reagent gave closely similar percentage values, the lymphocytes stained by the immunoenzymatic technique contrasted sharply from negative cells and were as a result more easily counted. Other practical advantages of immunoenzymatic staining over immunofluorescence are (1) the simultaneous visualization of cell morphology and type of label distribution over the cell surface, (2) cell scoring can be done rapidly on a large population sample, and (3) stained slides can be reexamined on subsequent days. A common advantages of Method SC for both immunofluorescence and immoenzymology is that antibody labeled slides can be stored before the final staining step.

Animals↗
PubMed1975

Two IgG1 (IgF) subclasses in mice: presence in normal serum and representation within lymphocyte and plasma cell populations.

Monospecific anti-gamma1 sera were prepared in rabbits against each of two IgG1 mouse myeloma globulins isolated from myeloma A2 and MOPC21. Both these sera react with IgG1 of normal mouse serum by immunoelectrophoresis and cannot be distinguished any further. The antibodies contained in the sera were cross-isolated on immunoadsorbents containing either the A2 or MOPC21 antigenic material and tested by immunoelectrophoresis against normal mouse serum. An interesting spur was formed between these two antibodies. The two isotypes thus revealed in normal serum are tentatively designated IgG1a (globulin A2) and Ig G1b (globulin MOPC21). With live peripheral lymph node cells almost all lymphocytes which stain with antikappa antibodies (B lymphocytes) also stain with the specific anti-gamma1a antibodies, but only 5-10% stain with the specific anti-gamma1b antibodies. On fixed spleen cells (C57B16 mice) the anti-gamma1a antibodies stain almost identical percentages of plasma cells as anti-kappa antibodies: 1.8% are gamma1 and 2.2% are kappa. Anti-gamma1b antibodies stain only 5% of kappa-positive plasma cells. The % values estimated on both the normal lymphocyte and plasma cell populations were not significantly affected by inhibtion experiments of the specific anti-gamma1 antibodies performed with isolated myeloma globulins representative of several Ig classes.

Animals↗