Document Detail

Calcium enhances the sensitivity of immunofluorescence for pemphigus antibodies.
MedLine Citation:
PMID:  3114387     Owner:  NLM     Status:  MEDLINE    
Indirect immunofluorescence (IF) to detect pemphigus and pemphigoid autoantibodies is commonly performed with monkey esophagus (ME) as substrate and phosphate-buffered saline (PBS) as a diluent. The purpose of this study was to evaluate comparative IF titers using human skin (HS) as substrate with variations in the buffers employed. Substrates (ME or HS) were incubated in PBS, Tris-acetate-buffered saline (TAS), TAS with 5 mM CaCl+2 (TAS-Ca+2), and PBS or TAS with 1 mM EDTA, prior to incubation with pemphigus or pemphigoid sera for indirect IF. We examined sera from 11 patients with pemphigus vulgaris (PV), 10 patients with Brazilian pemphigus foliaceus (BPF), and 4 patients with bullous pemphigoid. In 20 of 21 pemphigus sera, endpoint indirect IF titers were highest on normal skin with TAS-Ca+2. Six sera (2 PV and 4 BPF) had endpoints that were 5 double dilutions higher than the endpoints obtained with ME and PBS. Six sera (3 PV and 3 BPF) were 4 double dilutions higher, 7 sera (3 PV and 4 BPF) were 2-3 double dilutions higher, and 2 PV sera were equivalent with both substrate/buffers. Preincubation of either tissue with EDTA prior to indirect IF abolished PV and BPF antibody binding completely. Exposure to EDTA after the tissue was incubated with PV or BPF sera did not affect indirect IF titers. In the presence of Ca+2, the antigen was resistant to trypsin in concentrations of 0.001%; however, in the absence of added Ca+2 it was destroyed by 0.0001% trypsin. These differences were not observed with bullous pemphigoid sera; all 4 sera had similar endpoint indirect IF titers. This study shows a significant increase in the sensitivity of indirect IF assays for pemphigus autoantibodies by the use of Ca+2-supplemented buffers on human skin. This finding may also have implications for procedures designed to purify and/or detect pemphigus antigens.
W L Matis; G J Anhalt; L A Diaz; E A Rivitti; C R Martins; R S Berger
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  The Journal of investigative dermatology     Volume:  89     ISSN:  0022-202X     ISO Abbreviation:  J. Invest. Dermatol.     Publication Date:  1987 Sep 
Date Detail:
Created Date:  1987-10-20     Completed Date:  1987-10-20     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  0426720     Medline TA:  J Invest Dermatol     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  302-4     Citation Subset:  IM    
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MeSH Terms
Antibodies / analysis*
Calcium / pharmacology*
Dithiothreitol / pharmacology
Edetic Acid / pharmacology
Fluorescent Antibody Technique*
Infant, Newborn
Pemphigus / immunology*
Skin / immunology
Grant Support
Reg. No./Substance:
0/Antibodies; 3483-12-3/Dithiothreitol; 60-00-4/Edetic Acid; 7440-70-2/Calcium

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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