Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 2 de 2
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
MAbs ; 10(4): 539-546, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29485921

RESUMO

Monoclonal antibodies are commonly assumed to be monospecific, but anecdotal studies have reported genetic diversity in antibody heavy chain and light chain genes found within individual hybridomas. As the prevalence of such diversity has never been explored, we analyzed 185 random hybridomas, in a large multicenter dataset. The hybridomas analyzed were not biased towards those with cloning difficulties or known to have additional chains. Of the hybridomas we evaluated, 126 (68.1%) contained no additional productive chains, while the remaining 59 (31.9%) contained one or more additional productive heavy or light chains. The expression of additional chains degraded properties of the antibodies, including specificity, binding signal and/or signal-to-noise ratio, as determined by enzyme-linked immunosorbent assay and immunohistochemistry. The most abundant mRNA transcripts found in a hybridoma cell line did not necessarily encode the antibody chains providing the correct specificity. Consequently, when cloning antibody genes, functional validation of all possible VH and VL combinations is required to identify those with the highest affinity and lowest cross-reactivity. These findings, reflecting the current state of hybridomas used in research, reiterate the importance of using sequence-defined recombinant antibodies for research or diagnostic use.


Assuntos
Anticorpos Monoclonais/imunologia , Especificidade de Anticorpos/imunologia , Hibridomas/imunologia , Animais , Anticorpos Monoclonais/genética , Especificidade de Anticorpos/genética , Genes de Cadeia Pesada de Imunoglobulina/genética , Genes de Cadeia Pesada de Imunoglobulina/imunologia , Genes de Cadeia Leve de Imunoglobulina/genética , Genes de Cadeia Leve de Imunoglobulina/imunologia , Humanos
2.
Protein Eng Des Sel ; 29(1): 11-21, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26508747

RESUMO

The immunohistochemical (IHC) staining of mouse tissue sections using antibodies of mouse origin can result in high nonspecific background due to the staining of endogenous immunoglobulins (Igs) by enzyme-conjugated secondary antibodies. In order to obviate this issue, we developed a chimeric mouse-human anti-p53 monoclonal antibody (MH242) by grafting the variable regions of a known mouse antibody into a human Ig scaffold. This facilitated use of an anti-human secondary antibody, and resulted in near-zero background when compared with its parental mouse monoclonal antibody (PAb242). Furthermore, the chimeric antibody enabled reproducible detection of mutant p53 (homozygous R172H) expression in mouse tissue, an observation hitherto largely equivocal based on the use of existing antibodies. The approach we describe leads to the generation of tractable antibody reagents, whose integrity can be readily verified through DNA sequencing of expressor plasmids. The wide-spread adoption of such 'digitized' antibodies should reduce experimental disparities that can commonly arise through variations in antibody quality.


Assuntos
Anticorpos Monoclonais/química , Imuno-Histoquímica/métodos , Imagem Molecular/métodos , Proteínas Recombinantes de Fusão/química , Animais , Anticorpos Monoclonais/análise , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/metabolismo , Humanos , Imunoglobulina G/genética , Intestinos/química , Camundongos , Proteínas Recombinantes de Fusão/análise , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/imunologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA