V5多肽冻干粉

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Description

The V5 Peptide lyophilized powder was a synthetic peptide acquiring the amino acid residues N-Cys- Gly-Lys-Pro-Ile-Pro-Asn-Pro-Leu-Leu-Gly-Leu-Asp-Ser-Thr-C with a molecular weight of 1524.8, the core sequence stands for the same amino acid sequence for V5-tag (GKPIPNPLLGLDST) used in fusion protein expression.

Properties

It is highly suggested that the peptide powder is dissolved in buffer to make a 10x stock solution (like 4 mg/mL) and stored in -20°C in small size separately. Dilute into working concentration freshly before every elution operation. Do not store the working solution.

Application

The peptide can be used for competing out anti-V5 antibody binding to V5-tagged fusion proteins in immunoassays, thus it is a mild and ideal elution reagent for V5-tag fusion proteins from the anti-V5 monoclonal antibody in solution or bound to agarose on the Biotool Anti-V5 affinity gel.

Attention

1. Read the User Manual carefully before the first use;
2. Avoid freezing, drying and high-speed centrifugation during use and storage of beads;
3. This product is for R&D use only, not for drug, household, or other uses. Please consult the Material Safety Data Sheet for information regarding hazards and safe handling practices.

Procedure

Peptide Elution of V5 Fusion Protein from Biotool Anti-V5 affinity gel:
1. Thoroughly suspend the Anti-V5 Affinity Gel in the vial, for a uniform suspension of the resin. Quickly transfer 10μl of the gel suspension (about 5μl of packed gel volume) to a fresh tube.
2. Add 0.6ml TBS. Thoroughly suspend the Anti-V5 Affinity Gel by pipetting. Centrifuge the resin at 5000 rpm for 30 seconds and carefully remove the supernatant. Be sure to remove all of the wash buffer without discarding the resin. Repeat 3-4 times.
3. Add 500μl of cell periplasmic extracts to the washed resin.
4. Gently agitate samples for 2 hours at 4°C.
5. Centrifuge the resin for 30 seconds at 5000 rpm. Transfer the supernatants to a fresh tube.
6. Wash the resin with 0.5ml TBS until the OD280 of the supernatant reads＜0.05.
7. Elution of V5 Fusion Protein by Competition with V5 Peptide. Elute the bound V5 Fusion Protein by competitive elution with five one-column volume aliquots of a solution containing 100-400 ug/mL V5 Peptide in TBS.
8. Recycling the Biotool Anti-V5 affinity gel
V5 Peptide may not elute all of the V5 Fusion Protein bound to Biotool Anti-V5 affinity gel. It is recommended the Biotool Anti-V5 Affinity gel be regenerated immediately after use by washing with three 5 ml aliquots of 0.1 M glycine HCl, pH 3.5. The gel should be immediately re-equilibrated in TBS until the effluent is at neutral pH. 
Note: Do not leave the Biotool Anti-V5 Affinity gel in glycine HCl for longer than 20 minutes.
9. Storing the Biotool Anti-V5 Affinity gel
Wash the Biotool Anti-V5 Affinity gel three times with 5 ml of 50% glycerol with 10mM sodium phospV5te, 150mM sodium chloride, pH 7.4, containing 0.02% (w/v) sodium azide. then add another 5 ml of 50% glycerol with 10mM sodium phosphate, 150mM sodium chloride, pH 7.4, containing 0.02% (w/v) sodium azide and store at -20°C without draining.

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