A pre-embedding, protein a-gold immunolabelling technique for cytocentrifuged cell monolayers for lm and tem

1986 ◽  
Vol 44 ◽  
pp. 220-221
Author(s):  
K. Chien ◽  
I.P. Shintaku ◽  
A.F. Sassoon ◽  
R.L. Van de Velde ◽  
R. Heusser
Keyword(s):  
Cell Surface ◽  
Staining Method ◽  
Protein A ◽  
Surface Antigens ◽  
Cell Suspensions ◽  
Cellular Morphology ◽  
Cellular Phenotype ◽  
Cell Surface Antigens ◽  
Cell Monolayers ◽  
Diagnostic Histopathology

Identification of cellular phenotype by cell surface antigens in conjunction with ultrastructural analysis of cellular morphology can be a useful tool in the study of biologic processes as well as in diagnostic histopathology. In this abstract, we describe a simple pre-embedding, protein A-gold staining method which is designed for cell suspensions combining the handling convenience of slide-mounted cell monolayers and the ability to evaluate specimen staining specificity prior to EM embedding.

Immunogold staining method for the light microscopic detection of leukocyte cell surface antigens with monoclonal antibodies: its application to the enumeration of lymphocyte subpopulations.

1983 ◽  
Vol 31 (3) ◽  
pp. 376-381 ◽  
Author(s):  
M De Waele ◽  
J De Mey ◽  
M Moeremans ◽  
M De Brabander ◽  
B Van Camp
Keyword(s):  
Monoclonal Antibodies ◽  
Cell Surface ◽  
Peripheral Blood ◽  
Colloidal Gold ◽  
Staining Method ◽  
Surface Antigens ◽  
Lymphocyte Subpopulations ◽  
Cell Surface Antigens ◽  
Immunogold Staining ◽  
Microscopic Detection

Colloidal gold was used as a marker for the light microscopic detection of lymphocyte cell surface antigens with monoclonal antibodies. Suspensions of peripheral blood leukocytes were first incubated with monoclonal mouse antibodies and then with colloidal gold-labeled goat anti-mouse antibodies. The cells were fixed and cytocentrifuge preparations or smears were made. Granulocytes and monocytes were then labeled by the cytochemical staining of their endogenous peroxidase activity. Lymphocytes reacting with the monoclonal antibody had numerous dark granules around the surface membrane. With electron microscopy, these granules appeared as patches of gold particles. This immunogold staining method proved to be a reliable tool for the enumeration of T-lymphocyte subpopulations in peripheral blood. The results were almost identical to those obtained with immunofluorescence microscopy. The procedure can also be applied on small volumes of capillary blood. This constitutes a good microtechnique for the determination of lymphocyte subsets in children.

Indirect 125I-labeled protein A assay for monoclonal antibodies to cell surface antigens

1979 ◽  
Vol 31 (3-4) ◽  
pp. 201-209 ◽  
Author(s):  
Joseph P. Brown ◽  
John D. Tamerius ◽  
Ingegerd Hellström
Keyword(s):  
Monoclonal Antibodies ◽  
Cell Surface ◽  
Protein A ◽  
Surface Antigens ◽  
Cell Surface Antigens

scholarly journals An immunogold-silver staining method for detection of cell surface antigens in cell smears.

1989 ◽  
Vol 37 (12) ◽  
pp. 1855-1862 ◽  
Author(s):  
M De Waele ◽  
W Renmans ◽  
E Segers ◽  
V De Valck ◽  
K Jochmans ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Cell Surface ◽  
Silver Staining ◽  
Mononuclear Cells ◽  
Staining Method ◽  
General Procedure ◽  
Surface Antigens ◽  
Buffy Coat ◽  
Cell Surface Antigens ◽  
Silver Staining Method

We developed an indirect immunogold-silver staining method for detection of leukocyte cell surface antigens in cell smears. Air-dried and fixed cytocentrifuge preparations or smears of peripheral blood leukocytes were incubated with monoclonal antibodies (MAb) and colloidal gold-labeled secondary antibodies. The preparations were post-fixed and silver enhancement was performed. The smears were counterstained with May-Grunwald-Giemsa and examined in brightfield light microscopy. The morphology of the cells was well preserved. Leukocytes reacting with the MAb showed black granules on their surface membranes. The intense immunostaining and the low background allowed a rapid enumeration of the positive cells. The labeling could be detected with high sensitivity by epipolarization microscopy. This immunogold-silver staining method was used to quantify T- and B-lymphocytes and natural killer cells in buffy coat smears of normal adult blood. These lymphocyte subsets correlated well with those obtained in smears with the alkaline phosphatase-anti-alkaline phosphatase (APAAP) method and with those found by labeling of mononuclear cells in suspension with immunogold-silver staining. This immunogold-silver staining method forms a good alternative to immunoenzyme methods for study of hematologic cells. In addition, it could be a general procedure for detection of cell surface antigens in all kinds of cell smears.

scholarly journals An immunogold-silver staining method for detection of cell-surface antigens in light microscopy.

1986 ◽  
Vol 34 (7) ◽  
pp. 935-939 ◽  
Author(s):  
M De Waele ◽  
J De Mey ◽  
W Renmans ◽  
C Labeur ◽  
P Reynaert ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Cell Surface ◽  
Light Microscopy ◽  
Silver Staining ◽  
Surface Antigens ◽  
Cell Suspensions ◽  
T Cell Subsets ◽  
Staining Procedure ◽  
Cell Surface Antigens ◽  
Physical Developer

An immunogold-silver staining technique for detection of cell-surface antigens in cell suspensions was developed. Leukocyte cell suspensions were first incubated with monoclonal antibodies directed against cell-surface antigens and then with colloidal gold-labeled goat anti-mouse antibodies. Cytocentrifuge preparations of the cell suspensions were immersed in a physical developer containing silver lactate and hydroquinone as reducing substance. The preparations were then counterstained and mounted. In light microscopy, cells reacting with the monoclonal antibodies showed dark granules on their surface membrane. An optimal morphology, as revealed by a May-Grünwald-Giemsa counterstain, permitted accurate cell identification. The labeling was influenced by the gold particle diameter and the concentration of the gold reagents, by the duration of incubation in the physical developer, and by the composition and temperature of this medium. The T-cell subsets enumerated with this method in the peripheral blood of normal adults were identical to those found with other methods. The sensitivity of the technique was comparable with that of immunofluorescence microscopy. This immunogold-silver staining procedure proved to be a reliable tool for detection of cell-surface antigens in light microscopy.

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