Demonstration of cytoplasmic CD32 (Fc gamma RII) within human lymphocytes following microwave treatment.
We have recently described a cytoplasmic from of CD32 (Fc gamma RII) within the vast majority of normal human peripheral blood lymphocytes (PBL) including T cells. The function of cytoplasmic CD32 is not known. These flow cytometric studies were conducted using single cell suspensions of PBL that had been pre-fixed and permeabilized using methanol/triton-X-100. In this study we have attempted to visualize cytoplasmic CD32 by immunocytochemistry using normal PBL processed in various ways and have also looked for CD32 within tissue lymphocytes. Weak cytoplasmic CD32 staining was observed in paraffin sections of normal lymphocytes but only when sections were microwave treated. The intensity of staining for CD32 did however, appear to be much stronger within infiltrating lymphocytes found in autoimmune diseases or in rejecting allografts: an observation that suggests that up-regulation of cytoplasmic CD32 may occur when T cells become activated in vivo. Microwave treatment of PBL suspensions was shown to disrupt the outer cell membrane, thus effectively permeabilizing the cell and allowing for the detection of cytoplasmic components, like CD32, by flow cytometry. Microwave treatment may, therefore, afford an alternative method for cell permeabilization and may prove to be a useful method for the study of cytoplasmic molecules in cell suspensions and in paraffin-embedded tissues.
Sandilands
G P
GP
University Department of Pathology, Western Infirmary, Glasgow, UK.
Burnett
E R
ER
MacPherson
S A
SA
Downie
I
I
More
I A
IA
MacSween
R N
RN
D000911:Antibodies, Monoclonal; D002463:Cell Membrane Permeability / Q000528:radiation effects; D003593:Cytoplasm / Q000276:immunology*; D005434:Flow Cytometry; D006801:Humans; D007124:Immunoenzyme Techniques; D008214:Lymphocytes / Q000276:immunology* / Q000528:radiation effects / Q000648:ultrastructure; D008872:Microwaves; D016612:Paraffin Embedding; D017452:Receptors, IgG / Q000032:analysis*
D000911:Antibodies, Monoclonal; D017452:Receptors, IgG