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Zombie aqua fixable viability kit

Manufactured by Thermo Fisher Scientific
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Sourced in Belgium, United States

The Zombie Aqua Fixable Viability Kit is a lab equipment product designed to assess cell viability. It provides a method for distinguishing between live and dead cells in a sample.

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Lab products found in correlation

Lsrfortessa flow cytometer, by BD (2 mentions) Human fc block, by BD (1 mentions) Fixation solution, by BD (1 mentions) So lsrfortessa x20 flow cytometer, by BD (1 mentions) Cd10 pecf594, by BD (1 mentions) Clone 2.4g2, by BD (1 mentions) Siglec h clone 551, by BioLegend (1 mentions) Diva software, by BD (1 mentions) Pacific blue, by Thermo Fisher Scientific (1 mentions) Cd11b apc, by Thermo Fisher Scientific (1 mentions) Cd4 apc, by Thermo Fisher Scientific (1 mentions) Siglec f pe, by Thermo Fisher Scientific (1 mentions) Cd11b pacific blue, by BioLegend (1 mentions) F4 80 fitc, by Thermo Fisher Scientific (1 mentions) Dnase 1, by Roche (1 mentions) Cytoflex s, by Beckman Coulter (1 mentions) Apc anti mouse f4 80 clone bm8, by BD (1 mentions) Horizon fixable viability stain 700, by BD (1 mentions) Pe cy7 anti mouse ly6g, by BD (1 mentions) Fitc anti mouse ter 119, by BioLegend (1 mentions)

5 protocols using zombie aqua fixable viability kit

1

SARS-CoV-2 Spike Protein Binding Assay

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Two million of PBMCs were incubated with BD human FC block (BD Biosciences) and then stained with recombinant biotinylated spike S1 + S2 ECD-His (Sino Biological) conjugated with SA-R-Phycoerythrin (PE) and recombinant biotinylated-receptor binding domain  (RBD; BioLegend) conjugated with SA-Allophycocyanin (APC), together with the following fluorescent antibodies (clone; dilution used): CD3-BV650 (clone OKT3; 1:200); CD21-FITC (clone B-LY4; 1:50), CD19-BUV395 (clone SJ25C1; 1:100), CD10-PECF594 (clone HI10A; 1:200), IgM-BV605 (clone G20-127; 1:50), IgD-BV711 (clone IA6-2; 1:100), CD27-BV786 (clone O323; 1:50), CD11c-BB700 (clone 3.9; 1:50), CD20-APCH7 (clone 2H7; 1:100), CD38-BUV737 (clone HB7; 1:400), IgG-PE-Cy7 (clone G18-145; 1:100; all from Becton Dickinson), IgA-Vio blue (clone IS11-8E10; 1:100; Miltenyi Biotec). Following surface staining, cells were washed once with PBS and labeled with Zombie Aqua Fixable Viability Kit (Thermofisher) according to the manufacturer instruction. Cells were fixed in BD fixation solution (BD Biosciences) and acquired with SO LSRFortessa X20 flow cytometer (BD Biosciences). Data analysis was performed using FlowJo v10 (TreeStar, USA).
Polvere J., Fabbiani M., Pastore G., Rancan I., Rossetti B., Durante M., Zirpoli S., Morelli E., Pettini E., Lucchesi S., Fiorino F., Tumbarello M., Ciabattini A., Montagnani F, & Medaglini D. (2023). B cell response after SARS-CoV-2 mRNA vaccination in people living with HIV. Communications Medicine, 3, 13.
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2

Comprehensive T and B Cell Profiling

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Two million lung cells were incubated with Zombie Aqua™ Fixable Viability Kit (Thermo Fisher). After pre-incubation with an anti-CD16/CD32 antibody (Clone 2.4G2, BD Biosciences, Erembodegem, Belgium), cells were labelled with a combination of anti-mouse fluorochrome conjugated mAbs against CD3, CD4, CD8, CD25 and CD19 (BD Biosciences, Erembodegem, Belgium). Data acquisition was performed on a LSR Fortessa flow cytometer running DIVA software (BD Biosciences, Erembodegem, Belgium). FlowJo software was used for data analysis. Gating strategy for T and B cells are available in supplementary Figs. S3 and S4 respectively.
Decaesteker T., Jonckheere A.C., Vanhoffelen E., Schauvaerts J., Verhalle T., Cremer J., Dilissen E., Rodewald H.R., Dupont L., Bullens D.M.A, & Vanoirbeek J.A.J. (2022). Chlorine exposure and intensive exercise induces airway hyperreactivity in a 3-week murine exercise model. The Science of the total environment, 843.
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3

Lung Cell Phenotyping by Flow Cytometry

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Two million lung cells were incubated with Zombie Aqua™ Fixable Viability Kit (Thermo Fisher) and pre-incubated with an anti-CD16/CD32 antibody (Clone 2.4G2, BD Biosciences, Erembodegem, Belgium) to minimize nonspecific bindings. After live/dead staining, cells were labelled with combinations of anti-mouse fluorochrome-conjugated mAbs against CD45 (clone 30-F11), CD11c (clone N418), MHCII (clone M5/114.15.2), CD11b (clone M1/70), CD103 (clone 2E7), CD64 (clone X54-5/7.1) and Siglec-H (clone 551) (all from Biolegend, San Diego, California) (gating strategy see Fig. S5). Data acquisition was performed on a LSR Fortessa flow cytometer running DIVA software (BD Biosciences, Erembodegem, Belgium). FlowJo software (BD Bioscience, Ashland, Oregon) was used for data analysis.
Decaesteker T., Jonckheere A.C., Vanhoffelen E., Schauvaerts J., Verhalle T., Cremer J., Dilissen E., Rodewald H.R., Dupont L., Bullens D.M.A, & Vanoirbeek J.A.J. (2022). Chlorine exposure and intensive exercise induces airway hyperreactivity in a 3-week murine exercise model. The Science of the total environment, 843.
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4

Standardized Immune Cell Profiling

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Frozen PF cells were thawed in a 37°C water bath and immediately washed in PBS and 5% FBS. Cells were incubated with 100 μg/mL DNase 1 (Roche, 1014159001) at 4°C for 10 minutes. Cells were then passed through a 70 μm strainer to create a single-cell suspension. For surface marker staining, 5 × 105 cells/sample were incubated with the following monoclonal antibodies purchased from Biolegend, unless otherwise stated: Pacific blue–CD11b (M170), Superbright600-ST2 (RMST2-2), Bright Violet 785-CD45 (30-F11), FITC-F4/80 (BM8), PE-Siglec-F (E50-2440), PE/Cy7-FCER1 (MAR-1), APC-CD117 (2B8), Alexa Fluor 700-CD4 (RM4-5), APC/Cy7-MHC-II (M5/114.15.2), Pacific blue–CD90.2 (53.2.1), Brilliant Violet 785- CD45 (30-F11), PE-ST2 (Thermo Fisher Scientific, RMST2-2), FITC-CD25 (PC61), APC-CD3ε (145-2C11), APC-CD4 (RM4-5), APC-CD19 (1D3), APC-NK1.1 (PK136), APC-FCER1 (MAR-1), APC-Gr-1 (RB6-8C5), APC-CD11b (M1/70), Alexa Fluor 700–CD127 (Thermo Fisher Scientific, A7R34), and Zombie Aqua Fixable Viability Kit (no. 423101). Stained cells were analyzed on a Cytoflex S (Beckman Coulter), and the data were analyzed with FlowJo software (TreeStar). Live gate was set with one-half heat-killed viability control. Gates were set using FMOs. Full gating strategies are presented in Supplemental Figure 3.
Miller J.E., Lingegowda H., Symons L.K., Bougie O., Young S.L., Lessey B.A., Koti M, & Tayade C. (2021). IL-33 activates group 2 innate lymphoid cell expansion and modulates endometriosis. JCI Insight, 6(23), e149699.
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5

Multiparametric Flow Cytometry of Muscle Leukocytes

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Total leukocytes from naïve C57BL/6 muscle were isolated as described above and multiple populations identified using a CytoFLEX benchtop flow cytometer equipped with 405, 488, 561, and 637 nm solid phase lasers (Beckman Coulter Life Sciences, Brea, CA, USA) using the following fluorescent mAb (BioLegend):
FITC anti-mouse Ter119, PE/Cyanine7 anti-mouse Ly-6A/E (Sca-1), PE anti-mouse CD284 (TLR4), APC/Cy7 anti-mouse CD45, Brilliant Violet 421 anti-mouse CD31, and Zombie Aqua Fixable Viability Kit, as well as eFluor 660 anti-mouse/human CD34 (Thermo Fisher Scientific) (antibodies listed in Supplemental Table S2). At 4 days post-surgery, total leukocytes from muscle were isolated using a skeletal muscle dissociation kit (Miltenyi Biotech). Total leukocytes were subsequently discriminated into multiple populations using a CytoFLEX benchtop flow cytometer using the following mAb (BioLegend; outlined in Supplemental Table S1): FITC antimouse Ter119, FITC anti-mouse B220, FITC anti-mouse CD3ε, APC anti-mouse F4/80 (clone BM8), PE/Cy7 anti-mouse Ly6G, Brilliant Violet 510 anti-mouse/human CD11b, Brilliant Violet 785 anti-mouse CD45, and BD Horizon Fixable Viability Stain 700 (Supplemental Table S1). Files were subsequently analyzed with Flow Jo software version 10.5 (Becton Dickinson & Company, San Diego, CA, USA).
Salga M., Samuel S.G., Tseng H.W., Gatin L., Girard D., Rival B., Barbier V., Bisht K., Shatunova S., Debaud C., Winkler I.G., Paquereau J., Dinh A., Genêt G., Kerever S., Abback P.S., Banzet S., Genêt F., Lévesque J.P, & Alexander K.A. (2023). Bacterial Lipopolysaccharides Exacerbate Neurogenic Heterotopic Ossification Development. Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research, 38(11).
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