Immunoglobulin A in Graves' orbital tissue: deoxyribonucleic acid amplification by polymerase chain reaction.
A role for IgA autoantibodies in Graves' ophthalmopathy is suggested by the presence of immunoglobulins of this class in Graves' orbital tissue, as detected by immunohistochemistry. We, therefore, investigated the possibility of using the polymerase chain reaction (PCR) to amplify IgA immunoglobulin genes from plasma cells infiltrating Graves' eye tissue. Template cDNA was reverse-transcribed from orbital muscle (M) mRNA of one patient (#7) and from orbital connective tissue/fat (F) mRNA of two patients (#1 and #7), both undergoing surgery for exophthalmos because of severe infiltrative ophthalmopathy. Preliminary studies to establish the PCR procedure were performed for kappa light chain DNA amplification. With the very small amount of orbital tissue template available, the sensitive "hot start" modification of the PCR was necessary to amplify significant amounts of kappa light chain DNA. Using this procedure, IgA heavy chain DNA was amplified from both connective tissue/fat (F7) and muscle (M7) cDNA of patient #7. The DNA yield was less for IgA than for IgG using the same template. There was no significant IgA (or IgG) DNA product using the connective tissue/fat cDNA of patient #1. While not implying that IgA-infiltrating plasma cells are specific for Graves' orbital tissue, our studies nevertheless demonstrate the feasibility of amplifying the genes coding for IgA antibodies from Graves' orbital tissue plasma cells. Expression of these immunoglobulin genes in future studies will make it possible to determine the antigen specificity of the antibodies expressed by Graves' orbital tissue plasma cells.
McLachlan
S M
SM
Thyroid Molecular Biology Unit, V.A. Medical Center, San Francisco, California.
Prummel
M F
MF
Jaume
J C
JC
Rapoport
B
B
D000368:Aged; D004247:DNA / Q000235:genetics*; D005260:Female; D006111:Graves Disease / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007070:Immunoglobulin A / Q000032:analysis* / Q000235:genetics; D008297:Male; D008875:Middle Aged; D009915:Orbit / Q000737:chemistry*; D016133:Polymerase Chain Reaction / Q000379:methods
D007070:Immunoglobulin A; D004247:DNA