Intracellular staining of Mx proteins in cells from peripheral blood, bone marrow and skin.
OBJECTIVE
The Mx proteins are known to be specifically and dose dependently induced in mononuclear cells (MNC) by type I interferons (IFN). The aim of this study was to establish a staining method for the human intracellular Mx proteins, MxA and MxB, in leucocytes and bone marrow and skin cells.


METHODS
Several monoclonal antibodies directed against the MxA and MxB proteins were generated. These antibodies were used to stain Mx proteins in both frozen and paraffin wax sections using the standard alkaline phosphatase anti-alkaline phosphatase (APAAP) method.


RESULTS
Granulocytes, monocytes and lymphocytes extracted from freshly collected blood from 21 healthy subjects did not stain. After incubating MNC from these subjects with IFN alpha 2b for 48 hours, Mx proteins were detected in monocytes and lymphocytes. Within two days of starting treatment with subcutaneous IFN alpha 2b, granulocytes, monocytes and lymphocytes of 16 patients with cancer stained strongly for Mx proteins. The intensity of staining was correlated with the Mx content of whole blood measured using a specific ELISA. Prior to IFN treatment, cells from bone marrow and skin tissue specimens were negative for Mx proteins with the exception of endothelial cells. During treatment with IFN alpha 2b, nearly all cells from bone marrow and skin stained intensely.


CONCLUSIONS
These new monoclonal antibodies facilitate the detection of Mx positive cells in peripheral blood and in frozen or paraffin wax specimens. The advantage of this staining method is that individual cells which have responded to viruses or biologically active IFN alpha, beta or omega can be identified.
al-Masri
A N
AN
Department of Clinical Immunology, Medical School Hannover, Germany.
Werfel
T
T
Jakschies
D
D
von Wussow
P
P
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D000998:Antiviral Agents / Q000032:analysis* / Q000097:blood; D001853:Bone Marrow / Q000737:chemistry*; D005260:Female; D019204:GTP-Binding Proteins; D006801:Humans; D007124:Immunoenzyme Techniques; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000494:pharmacology; D008180:Lupus Erythematosus, Systemic / Q000097:blood; D051379:Mice; D008807:Mice, Inbred BALB C; D064547:Myxovirus Resistance Proteins; D011506:Proteins / Q000032:analysis*; D011994:Recombinant Proteins; D012867:Skin / Q000737:chemistry*; D013194:Staining and Labeling / Q000379:methods
D000911:Antibodies, Monoclonal; D000998:Antiviral Agents; D000077190:Interferon alpha-2; D016898:Interferon-alpha; C579071:MX1 protein, human; C579068:MX2 protein, human; C579070:Mx1 protein, mouse; D064547:Myxovirus Resistance Proteins; D011506:Proteins; D011994:Recombinant Proteins; D019204:GTP-Binding Proteins