Human PLA2G1B was produced in E. coli (gift of Gerard Lambeau, purity > 98%) or CHO-S (purity > 98%, Merck or GTP). Human proPLA2G1B, PLA2GIIA, PLA2GIID, PLA2GX, and WT and H48Q porcine PLA2G1B were produced in E. coli (gift of Gerard Lambeau, purity > 98%). Recombinant HIV-1 gp41 MN protein was obtained from Antibodies online (gp41 MN (565-771Delta642-725), containing a deletion to remove the transmembrane domain of the protein, ABIN2129703, lot 93-482, purity > 95%) and the 3S peptide NH2-PWNASWSNKSLDDIW-COOH, 3S-like peptide OmpA Pg NH2-SGEGGWSNGSLVDIM-COOH, and scrambled 3S NH2-WNWDSKILSDPAWNS-COOH peptide were ordered from Covalab (purity > 98%). The HCV core protein was obtained from Prospec (HCV-011, purity > 95%) in PBS buffer with 0.002% SDS and the specificity of the effect due to HCV core protein evaluated by comparison with a similar dilution of PBS-0.002% SDS. Staphylococcus aureus protein A was obtained from Sigma (P6031). sPLA2 inhibitors were obtained from several providers: Pentapeptide (sPLA2-IIA Inhibitor I, 525145, Calbiochem, ordered from VWR), Varespladib (LY315920, S1110, Selleck ordered from Euromedex), and sPLA2R1 (murine soluble receptor, 5367-PL-050, R&D Systems).
Staphylococcus aureus protein a
Manufactured by Merck Group
Sourced in United States
Staphylococcus aureus protein A is a cell wall-associated protein produced by the bacterium Staphylococcus aureus. It has the ability to bind to the Fc region of immunoglobulins, particularly IgG. This property makes it a useful tool in various laboratory applications, such as affinity chromatography and immunoassays.
Automatically generated - may contain errors
Lab products found in correlation
Ficoll 400, by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Trypsin, by Merck Group (2 mentions)
Poly l lysine (pll), by Merck Group (2 mentions)
Trypan blue, by Merck Group (2 mentions)
L glutamine solution, by Thermo Fisher Scientific (2 mentions)
Penicillin streptomycin solution, by Thermo Fisher Scientific (2 mentions)
Cho s, by Merck Group (1 mentions)
Varespladib, by Selleck Chemicals (1 mentions)
Cpg odn, by InvivoGen (1 mentions)
Pokeweed mitogen, by Merck Group (1 mentions)
Pcr purification kit, by Geneaid (1 mentions)
Tobramycin, by Merck Group (1 mentions)
Collagenase p, by Merck Group (1 mentions)
Mibefradil, by Alomone (1 mentions)
Alexa fluor 647 conjugated anti rabbit igg, by Jackson ImmunoResearch (1 mentions)
Anti vgcc rabbit polyclonal igg, by Alomone (1 mentions)
Hank s balanced salt solution, by Thermo Fisher Scientific (1 mentions)
Nickel 2 chloride, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
6 protocols using staphylococcus aureus protein a
1
Recombinant Phospholipase A2 Proteins
2
Quantifying Influenza HA-Specific IgG B Cells
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A/California/07/2009 (H1N1) influenza HA‐specific IgG‐secreting B cells were performed as previously described 21 . Briefly, PBMC were stimulated with a mixture of pokeweed mitogen (Sigma), Staphylococcus aureus protein A (Sigma) and CpG ODNs (Invivogen, San Diego, CA, USA) for 6 days before enzyme‐linked immunospot (ELISPOT) assay. H1N1 HA protein (Protein Sciences, Meriden, CT, USA) was used to coat the plate (Millipore, Watford, UK), and immunoglobulin (Ig)G‐ or IgA‐paired antibodies (Mabtech/Oxford lmmunotec Limited, Abingdon, UK) were used to reveal. ELISPOT readouts were expressed as the number of HA‐specific IgG or IgA ASC/106 PBMC.
3
ChIP Assay for Chromatin Immunoprecipitation
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The detailed procedure of the ChIP assay has been described in our previous papers [25 (link),28 (link),29 (link)]. Briefly, C2C12 cells were washed, fixed in formaldehyde (1%, w/v), and sonicated to shear chromatin. Specific antibody was added to the cleared lysate and the binding was allowed to proceed at 4°C overnight before Staphylococcus aureus Protein A (Sigma, #P7155) was added to capture the immune complex and then washed extensively. Then, the immune complex was eluted and the released DNA was extracted with phenol/chloroform twice and further purified using a PCR purification Kit (Geneaid). The primer sets used in the ChIP assay are listed in Supplementary Table S3.
4
Staphylococcus aureus Biofilm Disruption
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Pre-formed biofilms in chambered coverglass were also exposed to 10% SaF that had undergone various manipulations as described below. Staphylococcus aureus protein A was purchased from Sigma and used at a final concentration of 50 μg/mL. Biofilms treated with tobramycin (Sigma) were exposed for 24 h to 1000 µg/mL of the drug.
5
Isolation and Characterization of Cells
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Reagents were obtained from the following sources: collagenase P, bovine serum albumin (BSA), Ficoll 400, trypan blue, 4-(2-hydroxyethyl)-1 piperazineethanesulfonic acid (HEPES), poly-L-lysine, Staphylococcus aureus protein A, trypsin, insulin, guinea-pig complement, nickel chloride (II), and all salts of electrophysiological recordings from Sigma-Aldrich (St. Louis, MO, USA); mibefradil, nifedipine, anti-VGCC rabbit polyclonal IgG from Alomone Labs (Jerusalem, Israel); FITC-conjugated anti-guinea pig, and Alexa Fluor 647-conjugated anti-rabbit IgG from Jackson ImmunoResearch (West Grove, USA). Tetrodotoxin from Calbiochem (La Jolla, CA, USA); TTA-A2 kindly donated by Merck (West Point, PA, USA) to Dr. Juan Carlos Gomora; tissue culture dishes from Corning (Corning, NY, USA); Hanks’ balanced salt solution, RPMI-1640 salts, and penicillin–streptomycin solution, fetal bovine serum, and L-glutamine solution from Thermo Fisher Scientific Inc. (Massachusetts, USA).
6
Isolation and Culture of Primary Cells
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Reagents were obtained from the following sources: Hank's balanced salt solution (HBSS), ethylene glycolbis[β-aminoethylether]N,N,N′,N′-tetraacetic acid] (EGTA), Ficoll 400, trypan blue, poly-l-lysine, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), Staphylococcus aureus protein A, and trypsin from Sigma-Aldrich; collagenase P from Roche; DMEM salts, penicillinstreptomycin solution, Ham F10 medium, L-glutamine solution, fetal bovine serum (FBS), and guinea pig complement from Life Technologies; bovine serum albumin lyophilized powder (BSA) from United States Biological (Swampscott, MA, USA).
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