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. 2019 Jan 10;67(4):275–289. doi: 10.1369/0022155418824092

Figure 2.

Figure 2.

Effect of analyte concentration on ER immunostaining using the ER SP1 MAb, with and without HIER. Panels A and B depict immunostaining of peptide epitopes (coated onto glass microbeads) with (A) and without (B) HIER. These microbeads bear an average of 904,251 ER peptides per microbead. In Panel C, the ER peptide concentrations per microbead are listed above the vertical bars. The unstained test microbeads (labeled “Internal neg. ctrl microbead” in Panel B) bear an unrelated ER peptide epitope. They represent an internal negative control. The “Color standard microbead” (labeled in Panel B) is smaller and permanently brown, regardless of immunostaining. It is used as an internal optical reference standard for image quantification, normalizing variability in optical settings. Panels D and E depict immunostaining of the same peptide epitopes with (D) and without (E) HIER, except that these microbeads bear an average of 1331 peptide epitopes per microbead. The data (Panel C) represent the mean ± SD of triplicate slides. Scale bar, 10 µm. Abbreviations: ER, estrogen receptor; HIER, heat-induced epitope retrieval; PR, progesterone receptor.