Hantavirus antigen detection using human serum immunoglobulin M as the capturing antibody in an enzyme-linked immunosorbent assay.
An enzyme-linked immunosorbent assay (ELISA) was developed to detect different hantavirus antigens in cell culture; i.e. Puumala (PUU), Hantaan (HTN), and Dobrava (DOB) viruses. The assay was based on binding human serum immunoglobulin M (IgM) antibodies to the solid phase by use of goat anti-IgM antibodies. The captured IgM antibodies were present in the acute phase serum from two patients: one infected in Sweden and the other in Bosnia. Antigens being bound to the solid phase by the human anti-PUU and anti-DOB/HTN IgM antibodies were detected by a broadly reacting polyclonal rabbit anti PUU-recombinant nucleocapsid protein antiserum. The IgM isotype was proven to be at least five times more efficient than IgG when used as the capturing antibody. The sensitivity of the PUU antigen ELISA was approximately 0.5 ng/ml, as measured by titration with a PUU recombinant nucleoprotein antigen. Cell-associated PUU antigen in tissue culture was seen after 48 hr by the PUU-ELISA and after 96 hr by immunofluorescent assay. When tested for capacity to discriminate between PUU, DOB, and HTN viruses, significant differences were found: the Swedish serum detected PUU antigen at high titers, whereas no reactivity was found against DOB and HTN; the Bosnian serum detected both DOB and HTN at high titers but had a low reactivity to PUU. The method was also tested for its usefulness in detecting PUU antigen in bank vole (clethrionomys glareolus) lungs. Of 59 animals captured from the surroundings of patients with nephropathia epidemica, three became positive with a high activity in the PUU-ELISA, but with low reactivity in the DOB/HTN-ELISA. It is concluded that a sensitive ELISA has been developed to detect different hantaviruses in cell culture and lungs of bank voles.
Alexeyev
O A
OA
Department of Virology, University of Umea, Sweden.
Elgh
F
F
Ahlm
C
C
Stigbrand
T
T
Settergren
B
B
Wadell
G
G
Juto
P
P
D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology*; D000956:Antigens, Viral / Q000032:analysis*; D003411:Arvicolinae; D002522:Chlorocebus aethiops; D004306:Dose-Response Relationship, Immunologic; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D006041:Goats; D006476:Orthohantavirus / Q000276:immunology* / Q000302:isolation & purification; D018778:Hantavirus Infections / Q000175:diagnosis* / Q000276:immunology / Q000821:virology; D006480:Hemorrhagic Fever with Renal Syndrome / Q000276:immunology; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D007075:Immunoglobulin M / Q000276:immunology*; D008168:Lung / Q000821:virology; D011817:Rabbits; D051381:Rats; D012680:Sensitivity and Specificity; D014709:Vero Cells
D000914:Antibodies, Viral; D000956:Antigens, Viral; D007106:Immune Sera; D007074:Immunoglobulin G; D007075:Immunoglobulin M