A constructed HLA-A2-restricted pMAGE-A1(278-286) tetramer detects specific cytotoxic T lymphocytes in tumour tissues in situ.
OBJECTIVE
To construct a human leucocyte antigen (HLA)-A2-restricted peptide 278-286 of melanoma-associated antigen family A, 1 (pMAGE-A1(278-286)) tetramer to analyse the distribution of cytotoxic T lymphocytes (CTLs) in tumour tissue and tumour-adjacent normal tissue.


METHODS
A HLA-A2-pMAGE-A1(278-286) tetramer was constructed. The distribution of pMAGE-A1(278-286)-specific CTLs was investigated in tumour tissues and tumour-adjacent normal tissues from patients with hepatocellular carcinoma using in situ HLA-A2-pMAGE-A1(278-286) tetramer staining.


RESULTS
Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis indicated that HLA-A2 heavy and light chain proteins were successfully obtained. The successful construction of the HLA-A2-pMAGE-A1(278-286) monomer was confirmed with Western blot analysis using W6/32 antibody. Flow cytometry confirmed the specific binding of HLA-A2-pMAGE-A1(278-286) tetramer to pMAGE-A1(278-286)-specific CTLs. In situ HLA-A2-pMAGE-A1(278-286) tetramer staining demonstrated that the number of pMAGE-A1(278-286)-specific CTLs in tumour tissues was significantly higher than in tumour-adjacent normal tissues.


CONCLUSIONS
The HLA-A2-pMAGE-A1(278-286) tetramer was useful for the detection of pMAGE-A1(278-286)-specific CTLs in both tumour tissues and tumour-adjacent normal tissues. In situ tetramer staining is a powerful tool for investigating the distribution of pMAGE-A1278-286-specific CTLs in the tumour microenvironment.
Biological Targeting Diagnosis and Therapy Research Centre, Guangxi Medical University, Nanning, Guangxi, China.
Sodaly
S
Mongkhoune
Yu-An
YA
Xie
Yu-Qing
YQ
Wang
Yue
Y
Chen
Nuo
N
Zhou
Yi
Y
Peng
Jian
J
He
Xia
X
Yu
Su-Fang
SF
Zhou
Guo-Rong
GR
Luo
Yong-Xiang
YX
Zhao
Xiao-Ling
XL
Lu
D006528:Carcinoma, Hepatocellular / Q000276:immunology*; D005455:Fluorescent Antibody Technique; D015789:HLA-A2 Antigen / Q000276:immunology*; D006801:Humans; D008113:Liver Neoplasms / Q000276:immunology*; D018655:Lymphocyte Count / Q000379:methods; D009363:Neoplasm Proteins / Q000138:chemical synthesis / Q000276:immunology*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology*; D013194:Staining and Labeling / Q000379:methods; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014407:Tumor Cells, Cultured
D015789:HLA-A2 Antigen; C506530:MAGE-A1 protein (278-286), human; D009363:Neoplasm Proteins; D010446:Peptide Fragments
HLA-A2-pMAGE-A1278–286 tetramer; Tumour-specific cytotoxic T lymphocytes; in situ tetramer staining