Shrimp Hemocyanin (SHC) is a highly soluble hemocyanin derived from shrimp. It can be used as an alternative to Keyhole Limpet Hemocyanin (KLH) as a carrier protein for conjugation with low-molecular-weight molecules such as peptides, nucleic acids, drugs or toxins to render them highly immunogenic. Tests have shown that SHC exhibits immunogenicity comparable to KLH, while its solubility (especially after conjugation with haptens) is significantly higher than that of KLH, allowing greater flexibility in immunogen preparation protocols.Product Format: Lyophilized powder dissolved in PBS. Reconstitute with ultrapure water before use.
Product Features
High Immunogenicity: Perfect replacement for KLH;
High Conjugation Yield: Conjugation efficiency identical to KLH;
High Solubility: Significantly higher solubility than KLH, compatible with uncommon or poorly soluble haptens;
High Purity: Purity > 98%;
Excellent Performance: Validated in hundreds of antibody preparation cases;
Cost-Effective: Price much lower than KLH.
Usage
SHC Activation and Hapten Conjugation
SMCC Activation
1. Reconstitute lyophilized SHC (unactivated) with an appropriate volume of ultrapure water to a final concentration of 20 mg/mL;
2. Add 800 µL of freshly prepared SMCC solution (5 mg/mL in ultrapure water) to 1 mL of reconstituted SHC solution;
3. Incubate with gentle mixing for 60 min at room temperature or 30 min at 37 °C;
4. Remove excess SMCC by size-exclusion chromatography.
Conjugation with Hapten
1. Dissolve 20 mg of thiol-containing hapten in 5 mL of conjugation buffer (83 mM sodium phosphate, 0.1 M EDTA, 0.9 M NaCl, 0.1 mM TCEP, pH 7.2);
Note: 0.1 mM TCEP is optional; its main function is to reduce disulfide bonds in the hapten to thiol groups.
2. Immediately mix the hapten solution with activated SHC at a mass ratio of 1:0.6, and react for 2 h at room temperature or overnight at 4 °C;
Note: This conjugation ratio is recommended. Variations in hapten size and structure may affect conjugation efficiency. To ensure efficient conjugation, optimize the hapten-to-SHC ratio via multiple preliminary experiments.
3. Remove EDTA by size-exclusion chromatography.
EDC Activation and Hapten Conjugation
EDC Activation
1. Equilibrate EDC and NHS to room temperature before use;
2. Reconstitute lyophilized SHC (unactivated) with an appropriate volume of activation buffer (0.1 M MES, 0.5 M NaCl, pH 6.0) to a final concentration of 1 mg/mL;
3. Add 0.4 mg EDC and 0.6 mg NHS to 1 mL of the above SHC solution, mix well, and react for 15 min at room temperature;
4. Add 1.4 µL β-mercaptoethanol to quench EDC;
5. Remove excess EDC, NHS and β-mercaptoethanol by size-exclusion chromatography.
Conjugation with Hapten
1. Add an equimolar amount of hapten (dissolved in PBS) to the activated SHC solution, and react for 2 h at room temperature;
2. Add hydroxylamine to a final concentration of 10 mM to stop the reaction;
3. Remove excess hydroxylamine by size-exclusion chromatography.
Notes
1. This product is accurately quantified. Dissolve directly in the original vial at the desired concentration. DO NOT aliquot, as significant loss may occur due to product properties / static electricity;
2. The predicted extinction coefficient ε1% of SHC is 10. Adjust the coefficient when determining the concentration of SHC solution;
3. For your safety and health, wear a lab coat and disposable gloves during operation;
4. For research use only.
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