Published customer image: Image caption: . F16-4-4 recognizes a polymorphic determinant on MHC class Ia. The level of OX18 staining is ~30-fold higher intracellularly than extracellularly, which probably reflects this antibody's affinity for MHC class Ib antigens. (A) DA splenocytes were stained with different concentrations of fluorescent-labelled anti-MHC class Ia and Ib (clone OX18) or anti-MHC class Ia (F16-4-4) antibodies. For detection of intracellular MHC class I, antibody epitopes on the cell surface were first blocked with unlabeled antibody. Cells were then fixed, permeabilized and stained intracellularly with fluorescent labelled antibodies using the same volume as used for extracellular staining. Histograms show samples with comparable mean fluorescent intensity when stained with indicated antibody dilutions (in brackets) extracellularly (red) and intracellularly (blue). (B) F16-4-4 binds a paraformaldehyde (PFA) sensitive epitope on MHC class Ia proteins. Non-permeabilized splenocytes from DA and DA.1IR85 were incubated with saturating concentrations of F16-4-4 (upper panel) or OX18 (lower panel) before (+ PFA) or after (− PFA) incubation of cells in a 4% PFA solution. Note the decreased F16-4-4 staining of PFA- treated cells from DA.1IR85, which suggests that F16-4-4 binds a polymorphic determinant (similar PFA sensitive epitopes were found in DA.1HR83 and DA.1UR83 (data not shown)). Gray histograms represent fluorescence minus one (FMO) controls. From: Tuncel J, Haag S, Yau ACY, Norin U, Baud A, et al. (2014) Natural Polymorphisms in Tap2 Influence Negative Selection and CD4:CD8 Lineage Commitment in the Rat. PLoS Genet 10(2): e1004151 . This is from an open access article distributed under the terms of the Creative Commons Attribution License .