Detection of autoantibodies against alpha-2-macroglobulin-like 1 in paraneoplastic pemphigus sera utilizing novel green fluorescent protein-based immunoassays.
J Dermatol Sci
; 98(3): 173-178, 2020 Jun.
Article
em En
| MEDLINE
| ID: mdl-32439251
ABSTRACT
BACKGROUND:
Paraneoplastic pemphigus (PNP) is a devastating autoimmune multiorgan syndrome associated with autoantibodies against several autoantigens, including the alpha-2-macroglobulin-like-1 (A2ML1). A2ML1 is recognized by up to 70 % of PNP sera. The currently recommended techniques for serological diagnosis of PNP are inadequate to detect anti-A2ML1 antibodies.OBJECTIVES:
To develop novel assays which allow to easily and reliably detect anti-A2ML1 autoantibodies in PNP sera.METHODS:
We produced full-length A2ML1 in fusion with enhanced green fluorescent protein (EGFP-A2ML1) in transfected human embryonic kidney 293 T cells. The recombinant protein was used as fluorescent ligand for immunoprecipitation studies. We further developed an enzyme-linked immunosorbent assay (ELISA) by immobilizing EGFP-A2ML1 on 96-well plates.RESULTS:
A2ML1-positive PNP sera were able to immunoprecipitate EGFP-A2ML1. Direct measurement of fluorescence in immunoprecipitates correlates with the relative levels of anti-A2ML1 antibodies in the PNP sera. By the novel ELISA, based on the determined best cut-off value, 61 % of the tested 36 PNP sera were A2ML1 positive with a specificity of 88.9 % and a sensitivity of 95 %. The 20 tested normal sera (NHS) were negative, while 2 (10 %) of 20 pemphigus vulgaris and 3 (15 %) of 20 bullous pemphigoid sera showed borderline values.CONCLUSIONS:
Our novel immunoassays enable rapid stratification of PNP patients. The novel green fluorescent protein-based ELISA utilizing an active eukaryotic A2ML1 is highly sensitive and reliable and, hence, is useful for a better understanding of the immunological background of PNP. This approach may be easily applied for the rapid detection of antibodies to various other antigens.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Síndromes Paraneoplásicas
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Autoanticorpos
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Alfa-Macroglobulinas
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Penfigoide Bolhoso
/
Pênfigo
Tipo de estudo:
Diagnostic_studies
/
Evaluation_studies
Limite:
Humans
Idioma:
En
Revista:
J Dermatol Sci
Assunto da revista:
DERMATOLOGIA
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
Suíça