scholarly journals Multiparameter Flow Cytometry Assay for Analysis of Nitrosative Stress Status in Human Spermatozoa

2020 ◽  
Vol 97 (12) ◽  
pp. 1238-1247 ◽  
Author(s):  
Pamela Uribe ◽  
Juan Meriño ◽  
Emilio Manquemilla ◽  
Camila Villagrán ◽  
Etelinda Vega ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Nitrosative Stress ◽  
Human Spermatozoa ◽  
Multiparameter Flow Cytometry ◽  
Flow Cytometry Assay

10 A multiparameter flow cytometry assay to monitor natural killer cell proliferation and activation in immuno-oncology clinical trials

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A10-A10
Author(s):  
Jennifer Tsau ◽  
Brittney Atzmiller ◽  
David Quinn ◽  
Tanya Mulvey ◽  
Sema Kurtulus ◽  
...  
Keyword(s):  
Clinical Trial ◽  
Flow Cytometry ◽  
Cell Proliferation ◽  
T Cell ◽  
Natural Killer ◽  
Nk Cell ◽  
Multiparameter Flow Cytometry ◽  
Parent Population ◽  
Flow Cytometry Assay ◽  
Cd Markers

BackgroundNatural Killer (NK) cells have garnered increasing interest as potential cellular therapies or as targets of biotherapeutic agents due to their ability to kill tumor cells in a non-antigen dependent manner. Hence, measurement of NK cell proliferation and/or activation following treatment can serve as a useful biomarker for assessing the efficacy of immunomodulatory therapies.MethodsWe developed a novel 13-parameter flow cytometry panel incorporating cell differentiation (CD) markers important for identification of NK cell subsets (CD56, CD16), their proliferation (Ki-67), activation (CD25, CD335, NKG2D) and inhibition (CD159a) status. Additionally, CD markers that identify other cellular subsets known to be amenable to cytokine modulation (e.g., CD3 and CD14) were included for concurrent monitoring of T cell proliferation and monocyte activation. Method validation focused on analytical sensitivity, specificity and precision as key criteria of assay performance using peripheral blood mononuclear cells (PBMCs) stimulated with NK cell-activating cytokines and resting PBMCs from healthy donors.ResultsThe assay design allowed for robust quantitation of NK cell, T cell and monocyte functionalities. Lower limit of quantification (LLOQ) of target biomarker population was determined to be 1.0% of the parent population, based upon an analysis of 110 key target populations that displayed a co-efficient of variation (CV) of ≤25% and their frequencies ranged from 0.1% to 97.8% of the parent population. Additionally, ≤25% CV was observed in precision assessments, confirming the repeatability and reproducibility of the assay. Clinical trial utility of the assay was verified on cryopreserved PBMCs from patients with a variety of solid tumor malignancies. In these patients, the assay could clearly identify proliferating and activated NK cells, as well as proliferating T cells and activated monocytes, thus demonstrating its suitability for clinical trial applications.ConclusionsWe developed and validated a novel multiparameter flow cytometry assay that allows for simultaneous measurement of proliferation, activation and inhibitory status of key immune cell subsets. Thus, this assay can help shed light on the mode of efficacy of novel therapeutic agents that modulate the immune system, aimed at treatment of cancer and autoimmune diseases.

scholarly journals Next-generation multiparameter flow cytometry assay improves the assessment of oxidative stress in probiotics

2020 ◽  
Vol 91 ◽  
pp. 103501
Author(s):  
Vincenzo Fallico ◽  
Mary Rea ◽  
Catherine Stanton ◽  
Niclas Ilestam ◽  
Julie McKinney
Keyword(s):  
Oxidative Stress ◽  
Flow Cytometry ◽  
Multiparameter Flow Cytometry ◽  
Next Generation ◽  
Flow Cytometry Assay

Whole Spleen Flow Cytometry Assay

BIO-PROTOCOL ◽  
2013 ◽  
Vol 3 (15) ◽  
Author(s):  
Cathy Yam ◽  
Adeline Hajjar
Keyword(s):  
Flow Cytometry ◽  
Flow Cytometry Assay

scholarly journals Identification of pluripotent cells in bovine uterus: in situ and in vitro studies

Reproduction ◽  
2015 ◽  
Vol 149 (4) ◽  
pp. 317-327 ◽  
Author(s):  
Martyna Łupicka ◽  
Gabriel Bodek ◽  
Nahum Shpigel ◽  
Ehud Elnekave ◽  
Anna J Korzekwa
Keyword(s):  
Flow Cytometry ◽  
Protein Expression ◽  
Mrna Expression ◽  
Uterine Horn ◽  
Uterine Tissue ◽  
Pluripotent Cells ◽  
Flow Cytometry Assay ◽  
Myometrial Cells ◽  
Gene And Protein Expression

The aim of this study was to identify uterine pluripotent cells both in bovine uterine tissues as well in epithelial, stromal, and myometrial uterine cell populations. Moreover, the relationship of pluripotent markers expression with age and the uterine horn side was considered. Uterine tissue was collected from ipsilateral and contralateral horns (days 8–10 of the estrous cycle). Immunohistostaining for C-KIT, OCT3/4, NANOG, and SOX2 in uterine tissue was determined. mRNA expression of C-KIT, OCT3/4, NANOG and SOX2 was evaluated in uterine tissue relative to the age of the cow and uterine horn side. Gene and protein expression of these markers in the uterine luminal epithelial, stromal, and myometrial cells was evaluated by real-time PCR and western blotting respectively. The expression of pluripotent cell markers OCT3/4, NANOG, and SOX2 was identified by flow cytometry assay in epithelial, stromal, and myometrial cells. Multilineage differentiation of the bovine uterine cells was performed. mRNA expression of OCT3/4, NANOG, and SOX2 in uterine tissue was higher in the ipsilateral horn than in the contralateral horn. Flow cytometry assay revealed positive fluorescence for OCT3/4, NANOG, and SOX2 in all uterine cell types. Results showed the age-dependent expression of pluripotent markers in uterine tissue. Beside, the different expression of pluripotent cells in each horn of uterus suggests the influence of ovarian hormones on these characteristics. The highest mRNA and protein expression for pluripotent markers was observed in stromal cells among uterine cells, which indicates this population of cells as the main site of pluripotent cells in the cow uterus.

scholarly journals Assessment of Immunotoxicity by Multiparameter Flow Cytometry

1997 ◽  
Vol 38 (1) ◽  
pp. 38-54
Author(s):  
Scott W. Burchiel ◽  
Nancy L. Kerkvliet ◽  
G. Frank Gerberick ◽  
David A. Lawrence ◽  
Gregory S. Ladics
Keyword(s):  
Flow Cytometry ◽  
Multiparameter Flow Cytometry

scholarly journals Measurement of Separase Proteolytic Activity in Single Living Cells by a Fluorogenic Flow Cytometry Assay

PLoS ONE ◽  
2015 ◽  
Vol 10 (8) ◽  
pp. e0133769 ◽  
Author(s):  
Wiltrud Haaß ◽  
Helga Kleiner ◽  
Martin C. Müller ◽  
Wolf-Karsten Hofmann ◽  
Alice Fabarius ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Proteolytic Activity ◽  
Living Cells ◽  
Flow Cytometry Assay ◽  
Single Living Cells ◽  
Single Living
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