An immunohistochemical method for the detection of tumour necrosis factor alpha in cytospins of human bronchoalveolar lavage cells.
An immunohistochemical method for assessing the level of tumour necrosis factor-alpha in alveolar macrophages obtained by brochoalveolar lavage is described. Cytospins of mixed populations of lung cells were incubated first with a monoclonal antibody to CD68 and then with a specific peroxidase-labelled second antibody in a two-step reaction for the detection of the macrophage marker CD68. A second similarly based two-step reaction for the detection of tumour necrosis factor-alpha followed. Both reactions were visualized, on completion, using different coloured peroxidase substrates which produced a third colour in the event of dual deposition of the substrates. Dual substrate deposition was indicative of alveolar macrophages positive for tumour necrosis factor-alpha. This method has provided a specific and reproducible semi-quantitative test for the presence of tumour necrosis factor-alpha in human activated alveolar macrophages, which can be performed retrospectively on clinical material. A range of concentrations of the cytokine has been demonstrated in individual samples. This dual detection method has the potential for detection of any cell-associated protein product by minor modification of the described method.
Hamilton
S
S
Department of Virology, University of Newcastle Upon Tyne, Medical School, UK.
Healy
M
M
Corris
P
P
Scott
R
R
D000918:Antibody Specificity; D015703:Antigens, CD / Q000378:metabolism; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism; D001992:Bronchoalveolar Lavage Fluid / Q000166:cytology*; D002498:Centrifugation; D003584:Cytological Techniques; D006801:Humans; D007150:Immunohistochemistry; D016040:Lung Transplantation / Q000502:physiology; D016676:Macrophages, Alveolar / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D000097382:CD68 Molecule
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D014409:Tumor Necrosis Factor-alpha; C067980:CD68 antigen, human; D000097382:CD68 Molecule