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Mip 1β

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MIP-1β is a protein that functions as a chemokine, a type of signaling molecule that regulates the movement and activation of various immune cells. It is involved in the inflammatory response and plays a role in the recruitment and activation of specific leukocyte subpopulations.

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Lab products found in correlation

Ifn γ, by BD (7 mentions) Brefeldin a, by Merck Group (7 mentions) Tnf α, by BD (5 mentions) Golgistop, by BD (4 mentions) Staphylococcal enterotoxin b, by Merck Group (3 mentions) Golgistop, by Thermo Fisher Scientific (3 mentions) Anti cd107a antibody clone h4a3, by BD (3 mentions) Lsrii flow cytometer, by BD (3 mentions) Rosettesep, by STEMCELL (2 mentions) Cd49d l25, by BD (2 mentions) Cd103, by BD (2 mentions) Unlabeled cd28 l293, by BD (2 mentions) Eomesodermin wd1928 pe efluor610, by Thermo Fisher Scientific (2 mentions) S1pr1 sw4gypp efluor660, by Thermo Fisher Scientific (2 mentions) T bet, by BioLegend (2 mentions) Tnf α, by BioLegend (2 mentions) Interleukin 2 (il 2), by BioLegend (2 mentions) Clone b27, by BD (2 mentions) Fix perm, by Thermo Fisher Scientific (2 mentions) Lsr 2 flow cytometer, by BD (2 mentions)

Close spelling variants of this product

MIP-1β-PE-Cy7 (4 citations) APC-H7-conjugated MIP-1β mAbs (2 citations) PE–MIP-1β (2 citations) CCL4—MIP-1β (2 citations) Anti-MIP-1β-PerCP-Cy5.5 (2 citations)

15 protocols using mip 1β

1

Quantifying NK Cell Activation and Cytokine Production

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Human NK cells were enriched from human peripheral blood samples using negative selection with RosetteSep (Stem Cell Technologies) followed by Ficoll separation. Maxisorp ELISA plates were first coated with ZEBOV GPΔTM (3 μg/ml) before blocking with 5% BSA and incubation with 5 μg/ml mAbs at 37 °C for 2 hours. The mAbs were removed by washing and NK cells (5.0 × 104 cells/well) in the presence of 10 μg/ml brefeldin A (Sigma Aldrich), GolgiStop (BD), and anti-CD107a were incubated at 37 °C for 5 hr. The NK cells were then stained for NK surface markers (CD3, CD56, CD16, BD Biosciences) followed by intracellular staining to detect cytokine and chemokine production (IFN-γ and MIP-1β, BD Biosciences). NK cells were analyzed with a BD LSR2 flow cytometer and data were analyzed using FlowJo software.
Saphire E.O., Schendel S.L., Fusco M.L., Gangavarapu K., Gunn B.M., Wec A.Z., Halfmann P.J., Brannan J.M., Herbert A.S., Qiu X., Wagh K., He S., Giorgi E.E., Theiler J., Pommert K.B., Krause T.B., Turner H.L., Murin C.D., Pallesen J., Davidson E., Ahmed R., Aman M.J., Bukreyev A., Burton D.R., Crowe JE J.r., Davis C.W., Georgiou G., Krammer F., Kyratsous C.A., Lai J.R., Nykiforuk C., Pauly M.H., Rijal P., Takada A., Townsend A.R., Volchkov V., Walker L.M., Wang C.I., Zeitlin L., Doranz B.J., Ward A.B., Korber B., Kobinger G.P., Andersen K.G., Kawaoka Y., Alter G., Chandran K, & Dye J.M. (2018). Systematic analysis of monoclonal antibodies against Ebola virus GP defines features that contribute to protection. Cell, 174(4), 938-952.e13.
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2

Peptide-pulsed CD4+ T Cell Assay

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Peptide-pulsed autologous CD4+ T cells were used as antigen presenting cells (APC) for stimulation assays. APC were labeled with 5μM CellTrace Violet (CTV) as recommended by the manufacturer (Life Technologies, Carlsbad, CA) and pulsed with the Gag peptide for 30 m at room temperature and washed D-PBS and then resuspended in T-cell medium. 1 × 106 CD4+ T cells or T-cell clones were co-cultured with an equal amount of pulsed or unpulsed APC for 6 h with monensin (Golgi stop; BD Biosciences) and anti-human CD107a (H4A3; BD Biosciences). Cells were surface stained, fixed and permeabilized with Cytofix/Cytoperm solution (BD Biosciences) and stained for IFN-γ (#B27) and MIP1-β (# D21-1351) both from BD Biosciences.
Ayala V.I., Trivett M.T., Coren L.V., Jain S., Bohn P.S., Wiseman R.W., O’Connor D.H., Ohlen C, & Ott D.E. (2016). A Novel SIV Gag-Specific CD4+ T-Cell Clone Suppresses SIVmac239 Replication in CD4+ T Cells Revealing the Interplay between Antiviral Effector Cells and Their Infected Targets. Virology, 493, 100-112.
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3

Reagents and Antibodies for T-cell Culture

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AIM-V media and CD4+ and CD8+ Dynal beads were purchased from Invitrogen. RPMI-1640 media and X-Vivo 15 media (BioWhittaker™) were from Lonza Biologics, Inc. Pooled Human AB Serum (PHS) was from Valley Biomedical. Ficoll-Paque Premium was from GE Healthcare Sciences. Aldrithiol-2 (AT-2) and reduced L-Glutathione were from Sigma-Aldrich. Good manufacturing practice-certified (GMP-certified) Interleukin-2 (IL-2), GMP-certified Interleukin-4 (IL-4), Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF), GMP-certified Anti-CD3 antibody, CliniMACS CD4 and CD14 microbeads, CliniMACS LS Tubing sets, CryoMACS DMSO and CliniMACS PBS/EDTA buffer were from Miltenyi Biotec, Inc. Polyinosinic-polycytidylic acid – poly-L-lysine carboxymethylcellulose (Poly-ICLC) was from Oncovir, Inc. Fluorescent antibodies for IL-2, -IFNγ, -TNFα, -MIP-1β, -CD3, -CD4, and -CD8 were purchased from BD biosciences. Antibodies for CD86, -CD14, -CD11c, and -HLA-DR were purchased from Biolegend. Recombinant human IL-2 and IL-7 were from R&D Systems. The following antibodies were manufactured in-house at the AIDS and Cancer Virus Program, Leidos Biomedical Research, Inc., Frederick National Laboratory: anti-p24CA [94–0001], anti-p17MA [R193244], anti-gp120HIV [P3F5-D5-F8]. Anti-gp41TM(4E10) was obtained from Polymun [AB004].
MILLER E., SPADACCIA M., SABADO R., CHERTOVA E., BESS J J.r., Mac TRUBEY C., HOLMAN R.M., SALAZAR A., LIFSON J, & BHARDWAJ N. (2014). Autologous Aldrithiol-2-Inactivated HIV-1 Combined with Polyinosinic-polycytidylic acid–poly-L-lysine carboxymethylcellulose as a Vaccine Platform for Therapeutic Dendritic Cell Immunotherapy. Vaccine, 33(2), 388-395.
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4

Multiparametric Flow Cytometry Assay

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The following monoclonal antibodies were used in flow cytometry: CD8 (SK1: APC-H7), CCR7 (3D12: PE-Cy7), CD69 (L78: PE), IFN-γ (B27: PE-Cy7), MIP-1β (D21-1351: PerCP-Cy5.5), from BD Biosciences (San Jose, CA); CD103 (Ber-ACT8: Ax488), TNF-α (MAb11: BV605), IL-2 (MQ1-17H12: BV650), CD107a (H4A3: BV711), CD4 (RPA-TA: BV570, BV605), CD45RO (UCHL1: BV785), CD27 (O323: BV650), CD3 (UCHT-1: Ax700), and T-bet (4B10: BV711) from Biolegend (San Diego, CA). Unlabeled CD28 (L293), and CD49d (L25) were from BD Pharmingen (San Diego, CA). Eomesodermin (WD1928: PE-eFluor610) and S1PR1 (SW4GYPP: eFluor660) were from ThermoFisher (Waltham, MA). The HIV-1 Gag peptide pool (p55, HXB2 sequence) consisted of 15-mers with an 11 amino acid overlap (BD Biosciences). Staphylococcal enterotoxin B (SEB) was from Sigma Aldrich.
Kiniry B.E., Li S., Ganesh A., Hunt P.W., Somsouk M., Skinner P.J., Deeks S.G, & Shacklett B.L. (2017). Detection of HIV-1-Specific Gastrointestinal Tissue Resident CD8+ T-cells in Chronic Infection. Mucosal immunology, 11(3), 909-920.
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5

EBOV GPΔTM Antigen Binding Assay

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Recombinant EBOV GPΔTM antigen was coated onto a MaxiSorp 96-well plates (Nunc) at 300ng/well at 4°C overnight. Wells were washed with PBS and blocked with 5% BSA prior to addition of samples diluted 1:50 for 2h at 37°C. Unbound antibodies were removed by PBS wash, and NK cells enriched from the peripheral blood of human donors were added at 5×104 cells/well in the presence of 4μg/ml brefeldin A (Sigma Aldrich), 5μg/ml GolgiStop (Life Technologies) and anti-CD107a antibody (Clone H4A3, BD Biosciences) for 5h. Cells were fixed and permeabilized with Fix/Perm (Life Technologies) according to manufacturer’s instructions to stain for intracellular IFNγ (Clone B27, BD Biosciences) and MIP-1β (Clone D21–1351, BD Biosciences). Cells were analyzed on a BD LSRII flow cytometer.
Gunn B.M., McNamara R.P., Wood L., Taylor S., Devadhasan A., Guo W., Das J., Nilsson A., Shurtleff A., Dubey S., Eichberg M., Suscovich T.J., Saphire E.O., Lauffenburger D., Coller B.A., Simon J.K, & Alter G. (2023). Antibodies against the Ebola virus soluble glycoprotein are associated with long-term vaccine-mediated protection of non-human primates. Cell reports, 42(4), 112402.
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6

NK Cell Activation by MARV GP-specific Abs

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MARV GP-antigen was coated onto a MaxiSorp 96-well plate (Nunc) at 300 ng/well at 4°C overnight. Wells were washed with PBS and blocked with 5% bovine serum albumin (BSA) prior to addition of antibodies diluted to 10 μg/mL, or 5-fold serially diluted from 1 to 6.4 x 10−5 μg/mL, for 2 h at 37°C. Unbound antibodies were removed by PBS wash, and NK cells enriched from the peripheral blood of three different human donors were added at 5 x 104 cells/well in the presence of 4 μg/mL brefeldin A (MilliporeSigma), 5 μg/mL GolgiStop (Life Technologies) and anti-CD107a antibody (Clone H4A3, BD Biosciences) for 5 h. Cells were fixed and permeabilized with Fix/Perm (Life Technologies) according to the manufacturer’s instructions to stain for intracellular IFNγ (Clone B27, BD Biosciences) and MIP-1β (Clone D21-1351, BD Biosciences). Cells were analyzed on an LSRII flow cytometer (BD Biosciences).
Ilinykh P.A., Huang K., Santos R.I., Gilchuk P., Gunn B.M., Karim M.M., Liang J., Fouch M.E., Davidson E., Parekh D.V., Kimble J.B., Pietzsch C.A., Meyer M., Kuzmina N.A., Zeitlin L., Saphire E.O., Alter G., Crowe JE J.r, & Bukreyev A. (2020). NON-NEUTRALIZING ANTIBODIES FROM A MARBURG INFECTION SURVIVOR MEDIATE PROTECTION BY FC-EFFECTOR FUNCTIONS AND ENHANCING EFFICACY OF OTHER ANTIBODIES. Cell host & microbe, 27(6), 976-991.e11.
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7

Cytokine and Signaling Pathway Analysis

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RPMI-1640 medium, penicillin and streptomycin, PD098059 (ERK inhibitor), PP2 (LYN inhibitor), Brefeldin A (Golgi blocker) and TRI Reagent were from Sigma (St Louis, MO, USA). Fetal calf serum (FCS) was purchased from Gibco BRL (Grand Island, NY, USA) and ELISA Assay Kit of mouse IL-10, IL-12, TNF-α, IFN-γ, MIP-1α, MIP-1β and Rantes were from BD and TGF-β was from eBioscience. dNTPs, RevertAidTM M-MuLV Reverse Transcriptase, oligo dT, RNase inhibitor and other chemicals required for cDNA synthesis were from Fermentas (USA). Anti-phospho-H3 and Anti-acetyl-H3 Abs were obtained from Abcam and chromatin immunoprecipitation (ChIP) assay kits were purchased from Millipore (Bedford, MA, USA). GAPDH, phosphorylated and dephosphorylated form of Lyn and ERK-1/2 antibodies, CCR5 antibody were obtained from Santa Cruz Biotechnology (San Jose, CA, USA).
Das S., Banerjee S., Majumder S., Paul Chowdhury B., Goswami A., Halder K., Chakraborty U., Pal N.K, & Majumdar S. (2014). Immune Subversion by Mycobacterium tuberculosis through CCR5 Mediated Signaling: Involvement of IL-10. PLoS ONE, 9(4), e92477.
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8

RAVV GP-Specific Antibody and NK Cell Assay

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3 μg/ml of recombinant, trimeric RAVV GP was coated on a Maxisorp ELISA plate. Plates were blocked with 5% BSA prior to addition of antibodies MR191, MR78, or MR82 (range of concentration 5 μg/ml to 0.1 μg/ml) for 2 hours at 37 °C. Human NK cells were enriched from donor peripheral blood by negative selection using RosetteSep (Stem Cell Technologies) followed by ficoll separation and were incubated with IL-15 (1 ng/ml) overnight at 37°C. The antibodies were removed and the wells washed prior to addition of NK cells. The NK cells were added at 2.5 × 104 cells/well in the presence of brefeldin A (Sigma Aldrich), GolgiStop (BD), and anti-CD107a and incubated for 5 hours at 37°C. NK cells were stained for surface markers of NK cells (CD3, CD56, CD16, BD Biosciences), and then stained intracellularly for the production of cytokines and chemokines (IFNγ and MIP-1β, BD Biosciences). Cells were analyzed by flow cytometry on an LSR II flow cytometer (BD Biosciences), and the resulting data were analyzed using FlowJo software.
King L.B., Fusco M.L., Flyak A.I., Ilinykh P.A., Huang K., Gunn B., Kirchdoerfer R.N., Hastie K.M., Sangha A.K., Meiler J., Alter G., Bukreyev A., Crowe JE J.r, & Saphire E.O. (2018). The Marburgvirus-neutralizing human monoclonal antibody MR191 targets a conserved site to block virus receptor binding. Cell host & microbe, 23(1), 101-109.e4.
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9

Characterizing Engineered T Cell Responses

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CD27 (BioLegend Cat #302830) CCR7 (BioLegend Cat #353218) labeled antibodies were used to stain the expanded T cell populations as previously described.54 (link) Expression of the CD19 CARs was detected using the biotinylated F(ab′)2 fragment from goat anti-mouse immunoglobulin G (IgG) sera (specific for scFvs of murine origin; Jackson ImmunoResearch), followed by staining with streptavidin-PE (BD Biosciences/Pharmingen, Cat# 554061). Intracellular cytokine staining was performed as previously described.55 (link) Briefly, after T cells stopped expanding, CD3/28 beads were removed by magnetic separation. The following day, the T cells were mixed with K562s expressing CD19 at a ratio of 2:1 and incubated for 5 hr with GolgiStop (BD Biosciences). Surface CD4 (BioLegend Cat# 317433) and CD8 (BD Biosciences Cat# 560273) staining was performed, samples were fixed, and the following day, intracellular staining was performed as previously described,56 (link) with an LSR II flow cytometer (BD Biosciences, San Jose, CA) and FlowJo software (Tree Star, Ashland, OR). The following antibodies were used for intracellular staining: -IL-2 (BD Biosciences Cat# 554567), interferon γ (IFNγ) (BD Biosciences Cat# 552882, TNF (BD Biosciences Cat# 557647), and MIP-1β (BD Biosciences Cat# 560688).
Medvec A.R., Ecker C., Kong H., Winters E.A., Glover J., Varela-Rohena A, & Riley J.L. (2017). Improved Expansion and In Vivo Function of Patient T Cells by a Serum-free Medium. Molecular Therapy. Methods & Clinical Development, 8, 65-74.
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10

Ebola GP Antigen Antibody-Mediated NK Cell Activation

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Recombinant Ebola GP antigen (IBT Bioservices) was coated onto
MaxiSorp 96-well plates (Nunc) at 300 ng/well at 4 °C overnight.
Wells were washed with PBS and blocked with 5% BSA prior to addition of
antibodies (5 μg/ml) and incubation for 2 h at 37 °C. Unbound
antibodies were removed by washing with PBS, and NK cells enriched from the
peripheral blood of human donors were added at 5×104cells/well in the presence of 4 μg/ml brefeldin A (Sigma-Aldrich), 5
μg/ml GolgiStop (Life Technologies) and anti-CD107a antibody (Clone
H4A3, BD Biosciences) for 5 h. Cells were fixed and permeabilized with
Fix/Perm (Life Technologies) according to the manufacturer’s
instructions to stain for intracellular IFNγ (Clone B27, BD
Biosciences), and MIP-1β (Clone D21–1351, BD Biosciences).
Cells were analyzed on a BD LSRII flow cytometer. Gating strategy and
representative flow cytometry plots are shown in Figure S4.
Gunn B.M., Lu R., Slein M.D., Ilinykh P.A., Huang K., Atyeo C., Schendel S.L., Kim J., Cain C., Roy V., Suscovich T.J., Takada A., Halfmann P.J., Kawaoka Y., Pauthner M.G., Momo M., Goba A., Kanneh L., Andersen K.G., Schieffelin J.S., Grant D., Garry R.F., Saphire E.O., Bukreyev A, & Alter G. (2021). A Fc-Engineering approach to define functional humoral correlates of immunity against Ebola virus. Immunity, 54(4), 815-828.e5.
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