IntroductionMonitoring of innate myeloid cells (IMC) is broadly applied in basic and translational research, as well as in diagnostic patient care. Due to their immunophenotypic heterogeneity and... Show moreIntroductionMonitoring of innate myeloid cells (IMC) is broadly applied in basic and translational research, as well as in diagnostic patient care. Due to their immunophenotypic heterogeneity and biological plasticity, analysis of IMC populations typically requires large panels of markers. Currently, two cytometry-based techniques allow for the simultaneous detection of >= 40 markers: spectral flow cytometry (SFC) and mass cytometry (MC). However, little is known about the comparability of SFC and MC in studying IMC populations. MethodsWe evaluated the performance of two SFC and MC panels, which contained 21 common markers, for the identification and subsetting of blood IMC populations. Based on unsupervised clustering analysis, we systematically identified 24 leukocyte populations, including 21 IMC subsets, regardless of the cytometry technique. ResultsOverall, comparable results were observed between the two technologies regarding the relative distribution of these cell populations and the staining resolution of individual markers (Pearson's rho=0.99 and 0.55, respectively). However, minor differences were observed between the two techniques regarding intra-measurement variability (median coefficient of variation of 42.5% vs. 68.0% in SFC and MC, respectively; p<0.0001) and reproducibility, which were most likely due to the significantly longer acquisition times (median 16 min vs. 159 min) and lower recovery rates (median 53.1% vs. 26.8%) associated with SFC vs. MC. DiscussionAltogether, our results show a good correlation between SFC and MC for the identification, enumeration and characterization of IMC in blood, based on large panels (>20) of antibody reagents. Show less
Welters, M.J.P.; Santegoets, S.J.; Burg, S.H. van der 2020
Oropharyngeal squamous cell carcinoma (OPSCC) develops as a consequence of several mutations in the tumor suppressor pathways or after a progressive infection with high risk human papillomavirus ... Show moreOropharyngeal squamous cell carcinoma (OPSCC) develops as a consequence of several mutations in the tumor suppressor pathways or after a progressive infection with high risk human papillomavirus (HPV). The dismal side effects of the current standard of care and the clear involvement of the immune system has led to a surge in clinical trials that aim to reinforce the tumor-specific immune response as a new treatment option. In this review, we have focused on the most recent literature to discuss the new findings and insights on the role of different immune cells in the context of OPSCC and its etiology. We then applied this knowledge to describe potential biomarkers and analyzed the rationale and outcomes of earlier and ongoing immunotherapy trials. Finally, we describe new developments that are still at the preclinical phase and provide an outlook on what the near future may bring, now that several new and exciting techniques to study the immune system at the single cell level are being exploited. Show less
Checkpoint blockade immunotherapies have revolutionised cancer treatment in the last decade. Nevertheless, these are only beneficial for a small proportion of cancer patients. Important... Show moreCheckpoint blockade immunotherapies have revolutionised cancer treatment in the last decade. Nevertheless, these are only beneficial for a small proportion of cancer patients. Important prognosticators for response to immunotherapy are the mutation burden of tumours as well as the quality and quantity of tumour-infiltrating immune cells. High-throughput multiplex immunophenotyping technologies have a central role in deciphering the complexity of anti-tumour immune responses. Current techniques for the immunophenotyping of solid tumours are held back by the lack of spatial context, limitations in the number of targets that can be visualised simultaneously, and/or cumbersome protocols. We developed a tyramide signal amplification-free method for the simultaneous detection of seven cellular targets by immunofluorescence. This method overcomes limitations posed by most widespread techniques and provides a unique tool for extensive phenotyping by multispectral fluorescence microscopy. Furthermore, it can be easily implemented as a high-throughput technology for validation of discovery sets generated by RNA sequencing or mass cytometry and may serve in the future as a complementary diagnostic tool. Show less
Yazdi, M.T.; Schinkelshoek, M.S.; Loof, N.M.; Taube, C.; Hiemstra, P.S.; Welters, M.J.P.; Burg, S.H. van der 2016
