The synthetic peptide was cross-linked to a carrier protein keyhole limpet hemocyanin (KLH) using EDC (1-Ethyl-3-[dimethylaminopropyl] carbodiimide hydrochloride) and then injected into footpad of rabbits, five times with intervals of 2 weeks. At 11 weeks after the start of immunization, sera were obtained from the rabbits. The pooled antiserum was purified through a protein-A affinity column (nProtein A Sepharose 4 Fast Flow; GE Healthcare, code: 17528004) to extract the immunoglobulin fraction (conducted commercially by ARK Resource, Co., Ltd. Kumamoto, Japan). Subsequently, the fraction was purified by an Nhydroxy succinimide affinity column (NHS-activated Sepharose 4B; GE Healthcare Life Sciences, PA USA) coupled with GluN2B-NT2 or GluN1- NT peptides (prepared commercially by Green Peptide, Kurume, Japan). The fraction bound to NHS-activated Sepharose 4B gel column was eluted by glycine buffer (100 mM glycine, 0.5 M NaCl, pH 3.0).
Nhs activated sepharose 4b
Manufactured by GE Healthcare
Sourced in United States
NHS-activated Sepharose 4B is a ready-to-use affinity chromatography medium designed for the immobilization of ligands containing primary amino groups. It consists of highly cross-linked agarose beads with an N-hydroxysuccinimide (NHS) ester functional group, which can react with primary amines to form stable amide bonds. This product enables the simple and effective coupling of various biomolecules, such as proteins, enzymes, or peptides, for the development of affinity-based purification or detection methods.
Automatically generated - may contain errors
Lab products found in correlation
Nprotein a sepharose 4 fast flow, by GE Healthcare (1 mentions)
Chemical compounds, by Thermo Fisher Scientific (1 mentions)
Culture media, by Thermo Fisher Scientific (1 mentions)
Antibodies against e cadherin, by BD (1 mentions)
Hnrnpa2 b1, by Santa Cruz Biotechnology (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
β actin, by BD (1 mentions)
2 protocols using nhs activated sepharose 4b
1
Rabbit Antibody Production and Purification
2
Investigating AKT Signaling Inhibition
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Culture media, chemical
compounds, and fetal bovine serum (FBS) were
purchased from Life Technologies (Grand Island, NY, USA). Antibodies
against phospho-AKT (Ser473) and geldanamycin were purchased from
Cell Signaling (Temecula, CA, USA); antibodies against AKT, Slug,
and hnRNP A2/B1 were purchased from Santa Cruz Biotechnology (Santa
Cruz, CA, USA); antibodies against HSP90 and HSP70 were purchased
from Enzo Life Sciences (Farmingdale, NY, USA); and antibodies against
E-cadherin and β-actin were purchased from BD Sciences (San
Jose, CA, USA) and Sigma (St Louis, MO, USA), respectively. The HSP90
inhibitor 17AAG was purchased from Calbiochem (Gibbstown, NJ, USA).
The phenanthrene-based derivatives PBT-1 and PBT-1-6L (see Figure4 A for their structures) were synthesized in the
laboratory of Dr. K. H. Lee. NHS-activated Sepharose 4B was obtained
from GE Healthcare (Piscataway, NJ, USA). All oligonucleotides were
obtained from Bio Basic (Markham, ON, Canada).
compounds, and fetal bovine serum (FBS) were
purchased from Life Technologies (Grand Island, NY, USA). Antibodies
against phospho-AKT (Ser473) and geldanamycin were purchased from
Cell Signaling (Temecula, CA, USA); antibodies against AKT, Slug,
and hnRNP A2/B1 were purchased from Santa Cruz Biotechnology (Santa
Cruz, CA, USA); antibodies against HSP90 and HSP70 were purchased
from Enzo Life Sciences (Farmingdale, NY, USA); and antibodies against
E-cadherin and β-actin were purchased from BD Sciences (San
Jose, CA, USA) and Sigma (St Louis, MO, USA), respectively. The HSP90
inhibitor 17AAG was purchased from Calbiochem (Gibbstown, NJ, USA).
The phenanthrene-based derivatives PBT-1 and PBT-1-6L (see Figure
laboratory of Dr. K. H. Lee. NHS-activated Sepharose 4B was obtained
from GE Healthcare (Piscataway, NJ, USA). All oligonucleotides were
obtained from Bio Basic (Markham, ON, Canada).
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