Zombie uv live dead dye

Manufactured by BioLegend

Sourced in United States

Zombie UV Live/Dead dye is a fluorescent stain that can be used to distinguish between live and dead cells. It binds to DNA, with increased fluorescence upon binding to cells with compromised cell membranes, indicating cell death.

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Lab products found in correlation

Intracellular staining perm wash buffer, by BioLegend (6 mentions) Alexa fluor 488 donkey anti human igg, by Jackson ImmunoResearch (6 mentions) Fortessa x20, by BD (5 mentions) Facsaria 3, by BD (5 mentions) Lsrfortessa, by BD (4 mentions) Ic fix buffer, by Thermo Fisher Scientific (3 mentions) Cd27 pe cy7, by BioLegend (2 mentions) Cd86 bv711 it2, by BioLegend (1 mentions) Hla dr percpcy5.5 or pe cy7 l243, by BioLegend (1 mentions) Igm percp cy5, by BioLegend (1 mentions) Cd14 pedazzle594, by BioLegend (1 mentions) Igd pe, by BD (1 mentions) Cd3 buv737, by BD (1 mentions) Cd20 apc h7, by BD (1 mentions) Cd16 bv605, by BioLegend (1 mentions) Cd21 pe cf594, by BD (1 mentions) Cd16 bv650, by BioLegend (1 mentions) Cd3 bv650, by BD (1 mentions) Novoexpress 1, by Agilent Technologies (1 mentions) Novocyte, by Agilent Technologies (1 mentions)

Close spelling variants of this product

Live/Dead Zombie (4 citations) Zombie UV dye (29 citations) Live/Dead dye (5 citations) Zombie dye (26 citations) Zombie UV (60 citations)

14 protocols using zombie uv live dead dye

Flow Cytometric Analysis of SARS-CoV-2 Infection

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For flow cytometry analysis, adherent cells were recovered by trypsinising or gentle scraping and washed in PBS with 2 mM EDTA (PBS/EDTA). Non‐adherent cells were recovered from culture supernatants by centrifugation for 5 min at 300 g and washed once in PBS/EDTA. Cells were stained with fixable Zombie UV Live/Dead dye (BioLegend) for 6 min at room temperature. Excess stain was quenched with FBS‐complemented DMEM. For MDMs, Fc‐blocking was performed with PBS/EDTA+10% human serum for 10 min at 4°C. Cell surface with CD86‐Bv711 (IT2.2, BioLegend) and HLA‐DR‐PerCpCy5.5 or PE‐Cy7 (L243, BioLegend) staining was performed in PBS/EDTA at 4°C for 30min. Unbound antibody was washed off thoroughly, and cells were fixed in 4% PFA prior to intracellular staining. For intracellular detection of SARS‐CoV‐2 nucleoprotein, cells were permeabilised for 15 min with Intracellular Staining Perm Wash Buffer (BioLegend). Cells were then incubated with 1 μg/ml CR3009 SARS‐CoV‐2 cross‐reactive antibody (a kind gift from Dr. Laura McCoy) in permeabilisation buffer for 30 min at room temperature, washed once and incubated with secondary Alexa Fluor 488‐Donkey‐anti‐Human IgG (Jackson Labs). All samples were acquired on a BD Fortessa X20 or LSR II using BD FACSDiva software. Data were analysed using FlowJo v10 (Tree Star).

Thorne L.G., Reuschl A., Zuliani‐Alvarez L., Whelan M.V., Turner J., Noursadeghi M., Jolly C, & Towers G.J. (2021). SARS‐CoV‐2 sensing by RIG‐I and MDA5 links epithelial infection to macrophage inflammation. The EMBO Journal, 40(15), e107826.

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Phenotypic Analysis of Cryopreserved PBMCs

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Cryopreserved PBMCs were thawed and washed twice with 10 mL of FACS buffer (1 x PBS containing 2% FBS and 1 mM EDTA) and resuspended in 100 uL of 1x PBS containing Zombie UV live/dead dye at 1:200 dilution (BioLegend, 423108) and incubate at room temperature for 15 minutes. Following washing, cells were incubated with an antibody cocktail for 1 hour protected from light on ice. The following antibodies were used: IgD PE (BD Biosciences, 555779), IgM PerCP-Cy5.5 (BioLegend, 314512), CD20 APC-H7 (BD Biosciences, 560734), CD27 PE-Cy7 (BioLegend, 302838), CD14 PE/Dazzle^™ 594 (BioLegend, 301852), CD16 BV605 (BioLegend, 302040), IgG BV650 (BD Biosciences, 740596), CD3 BUV737 (BD Biosciences, 612750) and Alexa Fluor 488-labeled Wuhan spike (SinoBiological, 40589-V27B-B), and BV421 labeled Omicron Spike (SinoBiological^™, 40589-V49H3-B). All antibodies were used as the manufacturer’s instruction and the final concentration of each probe was 0.1 ug/ml. Cells were washed twice in FACS buffer and immediately acquired on a BD FACS Aria III for acquisition and FlowJo for analysis.

Wimmers F., Burrell A.R., Feng Y., Zheng H., Arunachalam P.S., Hu M., Spranger S., Nyhoff L., Joshi D., Trisal M., Awasthi M., Bellusci L., Ashraf U., Kowli S., Konvinse K.C., Yang E., Blanco M., Pellegrini K., Tharp G., Hagan T., Chinthrajah R.S., Grifoni A., Sette A., Nadeau K.C., Haslam D.B., Bosinger S.E., Wrammert J., Maecker H.T., Utz P.J., Wang T.T., Khurana S., Khatri P., Staat M.A, & Pulendran B. (2023). Systems biological assessment of the temporal dynamics of immunity to a viral infection in the first weeks and months of life. medRxiv.

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Flow Cytometric Detection of SARS-CoV-2 Nucleoprotein

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For flow cytometry analysis, adherent cells were recovered by trypsinization and washed in PBS with 2mM EDTA (PBS/EDTA). Cells were stained with fixable Zombie UV Live/Dead dye (Biolegend) for 6 min at room temperature. Excess stain was quenched with FBS-complemented DMEM. Unbound antibody was washed off thoroughly and cells were fixed in 4% PFA prior to intracellular staining. For intracellular detection of SARS-CoV-2 nucleoprotein, cells were permeabilized for 15 min with Intracellular Staining Perm Wash Buffer (BioLegend). Cells were then incubated with 1μg/ml CR3009 SARS-CoV-2 cross-reactive antibody (a kind gift from Dr. Laura McCoy) in permeabilization buffer for 30 min at room temperature, washed once and incubated with secondary Alexa Fluor 488-Donkey-anti-Human IgG (Jackson Labs). All samples were acquired on a BD Fortessa X20 using BD FACSDiva software. Data was analyzed using FlowJo v10 (Tree Star).

Reuschl A.K., Thorne L.G., Zuliani-Alvarez L., Bouhaddou M., Obernier K., Hiatt J., Soucheray M., Turner J., Fabius J.M., Nguyen G.T., Swaney D.L., Rosales R., White K.M., Avilés P., Kirby I.T., Melnyk J.E., Shi Y., Zhang Z., Shokat K.M., García-Sastre A., Jolly C., Towers G.J, & Krogan N.J. (2021). Host-directed therapies against early-lineage SARS-CoV-2 retain efficacy against B.1.1.7 variant. bioRxiv.

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Flow Cytometry for SARS-CoV-2 Nucleoprotein Detection

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For flow cytometry analysis, adherent cells were recovered by trypsinization and washed in PBS with 2 mM EDTA (PBS/EDTA). Cells were stained with fixable Zombie UV Live/Dead dye (BioLegend) for 6 min at room temperature. Excess stain was quenched with FBS-complemented DMEM. Unbound antibody was washed off thoroughly and cells were fixed in 4% PFA before intracellular staining. For intracellular detection of SARS-CoV-2 nucleoprotein, cells were permeabilized for 15 min with intracellular staining perm wash buffer (BioLegend). Cells were then incubated with 1 μg ml⁻¹ CR3009 SARS-CoV-2 cross-reactive antibody (a gift from L. McCoy) in permeabilization buffer for 30 min at room temperature, washed once and incubated with secondary Alexa Fluor 488-donkey-anti-human IgG (Jackson Labs). All samples were acquired on a BD Fortessa X20 using BD FACSDiva software. Data were analysed using FlowJo v.10 (Tree Star).

Thorne L.G., Bouhaddou M., Reuschl A.K., Zuliani-Alvarez L., Polacco B., Pelin A., Batra J., Whelan M.V., Hosmillo M., Fossati A., Ragazzini R., Jungreis I., Ummadi M., Rojc A., Turner J., Bischof M.L., Obernier K., Braberg H., Soucheray M., Richards A., Chen K.H., Harjai B., Memon D., Hiatt J., Rosales R., McGovern B.L., Jahun A., Fabius J.M., White K., Goodfellow I.G., Takeuchi Y., Bonfanti P., Shokat K., Jura N., Verba K., Noursadeghi M., Beltrao P., Kellis M., Swaney D.L., García-Sastre A., Jolly C., Towers G.J, & Krogan N.J. (2021). Evolution of enhanced innate immune evasion by SARS-CoV-2. Nature, 602(7897), 487-495.

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Intracellular SARS-CoV-2 Nucleoprotein Detection

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For flow cytometry analysis, adherent cells were recovered by trypsinisation and washed in PBS with 2mM EDTA (PBS/EDTA). Cells were stained with fixable Zombie UV Live/Dead dye (Biolegend) for 6 min at room temperature. Excess stain was quenched with FBS-complemented DMEM. Unbound antibody was washed off thoroughly and cells were fixed in 4% PFA prior to intracellular staining. For intracellular detection of SARS-CoV-2 nucleoprotein, cells were permeabilised for 15 min with Intracellular Staining Perm Wash Buffer (BioLegend). Cells were then incubated with 1μ/ml CR3009 SARS-CoV-2 cross-reactive antibody (a kind gift from Dr. Laura McCoy, UCL) in permeabilization buffer for 30 min at room temperature, washed once and incubated with secondary Alexa Fluor 488-Donkey-anti-Human IgG (Jackson Labs). All samples were acquired on a BD Fortessa X20 using BD FACSDiva software. Data was analysed using FlowJo v10 (Tree Star).

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Multiparametric Analysis of SARS-CoV-2 Antibody Profiles

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Cryopreserved PBMCs were thawed and washed twice with 10 ml of fluorescence-activated cell sorting (FACS) buffer (1× PBS containing 2% FBS and 1 mM EDTA) and resuspended in 100 μl of PBS containing Zombie UV LIVE/DEAD dye at 1:200 dilution (BioLegend, #423108) and incubated at room temperature for 15 min. After washing, cells were incubated with an antibody cocktail for 1 hour protected from light on ice. The following antibodies were used: IgD phycoerythrin (PE; Southern Biotech, #2030–09), IgM peridinin chlorophyll protein–Cy5.5 (BioLegend, #314512), CD20 allophycocyanin-H7 (BD Biosciences, #560734), CD27 PE-Cy7 (BioLegend, #302838), CD14 brilliant violet (BV) 650 (BioLegend, #301836), CD16 BV650 (BioLegend, #302042), IgG brilliant UV 395 (BD Biosciences, #564229), CD3 BV650 (BD Biosciences, #563916), CD21 PE-CF594 (BD Biosciences, #563474), Alexa Fluor 488–labeled Beta Spike protein (antibodies-online, #ABIN6963740), Alexa Fluor 647– labeled Omicron Spike protein (Sino Biological, #40589-V08H26), and BV421-l abeled Wuhan Spike protein (Sino Biological, #40589-V27B-B). All antibodies were used as per the manufacturer’s instruction, and the final concentration of each probe was 0.1 μg/ml. Cells were washed twice in FACS buffer and immediately acquired on a BD FACSAria III. FlowJo software v10 (TreeStar Inc.) was used for data analysis.

Arunachalam P.S., Feng Y., Ashraf U., Hu M., Walls A.C., Edara V.V., Zarnitsyna V.I., Aye P.P., Golden N., Miranda M.C., Green K.W., Threeton B.M., Maness N.J., Beddingfield B.J., Bohm R.P., Scheuermann S.E., Goff K., Dufour J., Russell-Lodrigue K., Kepl E., Fiala B., Wrenn S., Ravichandran R., Ellis D., Carter L., Rogers K., Shirreff L.M., Ferrell D.E., Deb Adhikary N.R., Fontenot J., Hammond H.L., Frieman M., Grifoni A., Sette A., O’Hagan D.T., Van Der Most R., Rappuoli R., Villinger F., Kleanthous H., Rappaport J., Suthar M.S., Veesler D., Wang T.T., King N.P, & Pulendran B. (2022). Durable protection against the SARS-CoV-2 Omicron variant is induced by an adjuvanted subunit vaccine. Science translational medicine, 14(658), eabq4130.

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SARS-CoV-2 Nucleocapsid Protein Quantification

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Infection levels were measured at 24 h by flow cytometry. Caco-2 cells were trypsinised, stained with fixable Zombie UV Live/Dead dye (BioLegend) and fixed with 4% PFA before intracellular staining for nucleocapsid protein. For intracellular detection of SARS-CoV-2 nucleoprotein, cells were permeabilised for 15 min with Intracellular Staining Perm Wash Buffer (BioLegend). Cells were then incubated with 1 μg/ml CR3009 SARS-CoV-2 cross-reactive antibody (a kind gift from Dr. Laura McCoy) in permeabilisation buffer for 30 min at room temperature, washed once and incubated with secondary Alexa Fluor 488-Donkey-anti-Human IgG (Jackson Labs). All samples were acquired and analysed using a NovoCyte (Agilent) and NovoExpress 1.5.0 software (Agilent). Supplementary Fig. 13 shows the gating strategy applied to a representative sample of Caco-2 cells.

Howell R., Clarke M.A., Reuschl A.K., Chen T., Abbott-Imboden S., Singer M., Lowe D.M., Bennett C.L., Chain B., Jolly C, & Fisher J. (2022). Executable network of SARS-CoV-2-host interaction predicts drug combination treatments. NPJ Digital Medicine, 5, 18.

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Modulation of B-T Cell Interaction

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Magnetic bead separation was used to isolate B cells (Mouse B Cell Isolation Kit; Miltenyi Biotec, Bergisch Gladbach, Germany) and CD4⁺ T cells (CD4⁺ T Cell Isolation kit; Miltenyi Biotec) from mechanically dissociated spleens from OTII mice. CD4⁺ T cells were labelled with CellTrace™ Violet Proliferation Dye (Thermo Fisher) and pre‐treated with titrating concentrations of BIIB091. Isolated B cells were pre‐treated with titrating concentrations of BIIB091 for 30 min at 37°C. To target OVA to BCR, BIIB091‐treated B cells were incubated with biotin‐conjugated goat F(ab′)2 anti‐mouse IgM (10 μg mL⁻¹; Jackson ImmunoResearch) on ice for 15 min, followed by washing and subsequent incubation with OVA‐conjugated anti‐biotin antibody (OVA Antigen Delivery Reagent; Miltenyi Biotec) for 10 min on ice. Titrating doses of BIIB091 were present during these two incubation steps. OVA‐targeted B cells were then cultured with CellTrace™ Violet‐labelled CD4 T cells in the presence of titrating concentrations of BIIB091 for 72 h at 37°C. Following the 72‐h incubation at 37°C, the B:T Co‐culture was labelled with Zombie UV™ live/dead dye (Biolegend) and stained for flow cytometry. The level of CD4⁺ T‐cell proliferation and the frequency of CD69⁺ B220⁺ B cells was quantified with FlowJo.

Bame E., Tang H., Burns J.C., Arefayene M., Michelsen K., Ma B., Marx I., Prince R., Roach A.M., Poreci U., Donaldson D., Cullen P., Casey F., Zhu J., Carlile T.M., Sangurdekar D., Zhang B., Trapa P., Santoro J., Muragan P., Pellerin A., Rubino S., Gianni D., Bajrami B., Peng X., Coppell A., Riester K., Belachew S., Mehta D., Palte M., Hopkins B.T., Scaramozza M., Franchimont N, & Mingueneau M. (2021). Next‐generation Bruton's tyrosine kinase inhibitor BIIB091 selectively and potently inhibits B cell and Fc receptor signaling and downstream functions in B cells and myeloid cells. Clinical & Translational Immunology, 10(6), e1295.

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SARS-CoV-2 Nucleoprotein Detection by Flow Cytometry

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For flow cytometry analysis, adherent cells were recovered by trypsinisation and washed in PBS with 2mM EDTA (PBS/EDTA). Cells were stained with fixable Zombie UV Live/Dead dye (Biolegend) for 6 min at room temperature. Excess stain was quenched with FBS-complemented DMEM. Unbound antibody was washed off thoroughly and cells were fixed in 4% PFA prior to intracellular staining. For intracellular detection of SARS-CoV-2 nucleoprotein, cells were permeabilised for 15 min with Intracellular Staining Perm Wash Buffer (BioLegend). Cells were then incubated with 1μg/ml CR3009 SARS-CoV-2 cross-reactive antibody (a kind gift from Dr. Laura McCoy, UCL) in permeabilization buffer for 30 min at room temperature, washed once and incubated with secondary Alexa Fluor 488-Donkey-anti-Human IgG (Jackson Labs). All samples were acquired on a BD Fortessa X20 using BD FACSDiva software. Data was analysed using FlowJo v10 (Tree Star).

Thorne L.G., Bouhaddou M., Reuschl A.K., Zuliani-Alvarez L., Polacco B., Pelin A., Batra J., Whelan M.V., Ummadi M., Rojc A., Turner J., Obernier K., Braberg H., Soucheray M., Richards A., Chen K.H., Harjai B., Memon D., Hosmillo M., Hiatt J., Jahun A., Goodfellow I.G., Fabius J.M., Shokat K., Jura N., Verba K., Noursadeghi M., Beltrao P., Swaney D.L., Garcia-Sastre A., Jolly C., Towers G.J, & Krogan N.J. (2021). Evolution of enhanced innate immune evasion by the SARS-CoV-2 B.1.1.7 UK variant. bioRxiv.

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Isolation and Characterization of Uterine Immune Cells

Cited 1 time

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Uterine samples consisting of maternal decidua and myometrium were
collected 6 h after ultrapure LPS challenge (or saline) on d16 of gestation
from WT and BAFF reporter (RFP^{48 (link)}) mice. Immune cells were isolated using enzymatic
digestion. Briefly, uterine tissue was dissected and finely minced. Tissue
was digested using Liberase TM (21 µg/mL, Roche) and Dnase I (8.8
µg/mL, Roche) in DMEM containing HEPES and 2% BSA. Tissue was
incubated at 37°C for 30 min at 220 RPM. After digestion, cells were
filtered through a 100 µM strainer and centrifuged at 800 g for 5
min. Following red blood cell lysis, single cell suspensions were stained
with Live/Dead (Zombie UV Dye: Biolegend) and directly conjugated monoclonal
antibodies to CD45-PE-Dazzle594 (Biolegend, 104), F4/80-AF700 (Biolegend,
BM8), TCRb-APC-ef780 (Invitrogen, H57–597), CD11b-ef450 (Biolegend,
17A2), CD8-BV510 (Biolegend, 53–6.7), B220-BV605 (Biolegend,
RA3–6B2), NK1.1-FITC (Biolegend, PK136), CD11c-BV711 (Biolegend,
N418), Ly6G-APC (Invitrogen, RB6–8C5), Ly6C-PerCP-Cy5.5 (eBioscience,
HK1.4). Data was collected using LSR Fortessa (BD) and analyzed using FlowJo
X software (vX10).

Smith H.C., Kuo S.R., Backus J.W., Harris S.G., Sparks C.E, & Sparks J.D. (1991). In vitro apolipoprotein B mRNA editing: identification of a 27S editing complex.. Proceedings of the National Academy of Sciences of the United States of America, 88(4), 1489-1493.

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