No abstract

Immune-profiling is becoming an important tool to identify predictive markers for the response to immunotherapy. Our goal was to validate multiplex immunofluorescence (mIF) panels to apply to formalin-fixed and paraffin-embedded tissues using a set of immune marker antibodies, with the Opal™ 7 color Kit (PerkinElmer) in the same tissue section. We validated and we described two panels aiming to characterize the expression of PD-L1, PD-1, and subsets of tumor associated immune cells. Panel 1 included pancytokeratin (AE1/AE3), PD-L1, CD4, CD8, CD3, CD68, and DAPI, and Panel 2 included pancytokeratin, PD-1, CD45RO, granzyme B, CD57, FOXP3, and DAPI. After all primary antibodies were tested in positive and negative controls by immunohistochemistry and uniplex IF, panels were developed and simultaneous marker expressions were quantified using the Vectra 3.0™ multispectral microscopy and image analysis InForm™ 2.2.1 software (PerkinElmer).These two mIF panels demonstrated specific co-localization in different cells that can identify the expression of PD-L1 in malignant cells and macrophages, and different T-cell subpopulations. This mIF methodology can be an invaluable tool for tumor tissue immune-profiling to allow multiple targets in the same tissue section and we provide that is accurate and reproducible method when is performed carefully under pathologist supervision.

KSHV strains in East and West Africa are closely related but phylogenetically distant from those in Europe. The two major KSHV groups in Europe are more closely related, with some strains adopting an intermediate phylogenetic position. In Europe, KSHV strains may have been disseminated at least several decades ago. Variability in the K1 region is driven by selection and does not correlate with different KSHV-related pathologies or geographic regions where clinically more aggressive HIV-negative KS ('endemic' KS) is more common.

Background and Purpose-Several studies have implied an association between Chlamydia pneumoniae and atherosclerosis. Our research was designed to investigate the association of this organism with strokes and transient cerebral ischemia. Methods-Antibodies to C pneumoniae were measured in 176 patients with stroke or transient cerebral ischemia and 1518 control subjects with noncardiovascular, nonpulmonary disorders. Acute infection or reinfection was defined by IgG Ն512 or IgM Ն8 or fourfold rise in IgG, and previous infection was defined by IgG 64 to 256 or lgA Ն8. Logistic regression was used to examine the influences of ethnic origin, age, sex, smoking habit, diabetes mellitus, steroid medication, and social deprivation on antibody levels. Some patients underwent CT and carotid ultrasound examinations and cholesterol, triglyceride, fibrinogen, and von Willebrand factor estimations. Results-We found that 13.6% of stroke/transient ischemic attack (TIA) patients and 5.7% of control subjects had antibody titers suggesting acute C pneumoniae (re)infection, while 32.4% of stroke/TIA patients and 12.7% of control subjects had titers suggesting previous infection (PϽ.05). Stroke/TIA patients differed from control subjects in their levels of acute and previous infection, with adjusted odds ratios of 4.2 (95% CI, 2.5 to 7.1) and 4.4 (95% CI, 3.0 to 6.5), respectively. These did not differ notably between strokes resulting from major nonhemorrhagic infarcts, small-vessel infarcts, or hemorrhage. Cholesterol, triglyceride, fibrinogen, and von Willebrand factor concentrations showed no apparent association with titers. Conclusions-These data support the association of cerebral vascular disease with previous C pneumoniae infection and the association of stroke and transient cerebral ischemia with recrudescence of infection. (Stroke. 1998;29:404-410.)