Fig 1.
Flow cytometry gating strategy for analyzing HLA cell surface expression on HEK293T cells.
Data are representative for all flow cytometry experiments. (A) Forward scatter vs. side scatter dot plot. (B) Histogram showing cell surface expression of HLA-A3 and -B7 using allele-specific antibodies either unstimulated or after IFN-γ stimulation.
Fig 2.
Allele-specific HLA class I cell surface expression and gene transcript levels of HEK293T cells.
Horizontal lines indicate median values. (A) Quantitative measurements of HLA class I cell surface expression determined by indirect flow cytometry. (B) HLA class I gene transcripts levels determined by quantitative real-time PCR. * Indicates significant P value (P < 0.05).
Fig 3.
Alignment of amino acid sequences deduced from HLA-A*02:01:01:01 and HLA-B*08:01:01.
Identical amino acids are shown as dots. Arrows indicate sites of transition between chimeric constructs. GenBank accession numbers for HLA-A*02:01:01:01, HG794376.1 and for HLA-B*08:01:01, HG794374.1 (http://www.ncbi.nlm.nih.gov/genbank/).
Fig 4.
Allele-specific HLA class I cell surface expression and gene transcript levels in cloned HEK293T cells transfected with either A2 or B8 coding sequence constructs.
(A) Quantitative measurement performed by indirect flow cytometry of HLA class I cell surface expression. Horizontal lines indicate median values. * Indicates significant P value (P < 0.05). (B) Cell surface protein expression of A2 and B8 as a function of mRNA copy number relative to reference gene copy number.
Fig 5.
Allele-specific HLA class I cell surface expression levels of cloned HEK293T cells transfected with chimeric constructs.
Quantitative measurement determined by indirect flow cytometry, horizontal lines indicate median values. (A) Cells transfected with either B8 or chimeric constructs starting N-terminally as B8 and ending C-terminally as A2. (B) Cells transfected with either A2 or chimeric constructs starting N-terminally as A2 and ending C-terminally as B8. * Indicates significant P value after Bonferroni correction (P < 0.017).