Phospho lck

Manufactured by BD

Phospho-LCK is a laboratory reagent used in the detection and quantification of the phosphorylated form of the LCK (Lymphocyte-Specific Protein Tyrosine Kinase) protein. LCK is an important signaling molecule in T cell activation and development. The Phospho-LCK reagent can be used in various analytical techniques, such as Western blotting and ELISA, to measure the levels of phosphorylated LCK in biological samples.

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Lab products found in correlation

Facsverse flow cytometer, by BD (1 mentions) Fcs express, by BD (1 mentions) Cd160, by BD (1 mentions) Phospho erk1 2, by BD (1 mentions) Dnam 1, by BioLegend (1 mentions)

2 protocols using phospho lck

Phosphoflow Analysis of PBMC Signaling

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PBMCs from healthy patients were treated with each drug for 2 hours and stimulated with 3.3 mM H₂O₂ for 10 minutes at 37°C. The cells were then fixed with 1.6% paraformaldehyde solution for 10 minutes at 37°C and permeabilized with 100% methanol overnight at –80°C. Cells were stained with antibodies against phospho-LCK (#558552; BD Biosciences, Franklin Lakes, NJ), phospho-SRC (#560096; BD Biosciences), cleaved Poly(ADP-ribose) polymerase (PARP) (#560640; BD Biosciences) and CD3 (#562426; BD Biosciences). A similar assay was previously described for T-cell phosphoflow analysis using CD3/CD28 and H₂O₂ stimuli (38 (link),39 (link)). Data were acquired on a FACSVerse flow cytometer and analyzed with FCS Express (version 4; BD Biosciences).

Patel V., Balakrishnan K., Bibikova E., Ayres M., Keating M.J., Wierda W.G, & Gandhi V. (2016). Comparison of acalabrutinib, a selective Bruton tyrosine kinase inhibitor, with ibrutinib in chronic lymphocytic leukemia cells. Clinical cancer research : an official journal of the American Association for Cancer Research, 23(14), 3734-3743.

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Multiparametric Flow Cytometry Assay

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For flow cytometry, fluorochrome-conjugated antibodies against the following antigens were used: CD56 (clone NCAM16.2; conjugated to BV605) (B159; APC), CD3 (SK7; BUV395) (HIT3a; PE), CD107a (H4A3; APC), IFN-γ (B27; PerCP/Cy5.5), TNF (MAb11; PE), CD69 (FN50; BV421 and BV711), CD11a (HI111; BV711), NKG2D (149810; PE-CF594), NKp30 (p30-15; BV421), TACTILE (6F9; BV711), TRAIL (RIK-2; BV421), BTLA (J168-540; PE-CF594), Phospho-Lck (4/LCK-Y505; AF647), Phospho-LAT (J96-1238.58.93; PE), Phospho-SLP-76 (J141-668.36.58; PE), Phospho-MEK1/2 (O24-836; PE), Phospho-ERK1/2 (20A; BV421), Phospho-SHP-2 (L99-921; AF647) (all BD Biosciences), CD160 (BY55; PE/Cy7), NKp46 (9E2; PE/Cy7),NKp44 (P44-8; PE), DNAM-1 (11A8; APC), 2B4 (C1.7; PerCP/Cy5.5), NKp80 (5D12; PE) (all BioLegend), NKG2C (134591; APC) (R&D Systems), and VZV gE:gI (SG1-1; conjugated in-house to DyLight 488) (Meridian Life Science). Matched isotype control antibodies were also used where appropriate.

Campbell T.M., McSharry B.P., Steain M., Russell T.A., Tscharke D.C., Kennedy J.J., Slobedman B, & Abendroth A. (2019). Functional paralysis of human natural killer cells by alphaherpesviruses. PLoS Pathogens, 15(6), e1007784.

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