- Производитель:
- Roche
Enzyme Commission (EC) Number: | 1.11.1.7 ( BRENDA | IUBMB ) |
Donor: hydrogen-peroxide oxidoreductase
Reagent for the labeling of water-soluble substances carrying primary amino groups with peroxidase from horseradish (HRP).
Capacity: sufficient for approximately 6 mg IgG.
Peroxidase (POD), activated has been used in terminal deoxyribonucleotidyl transferase (TDT)-mediated dUTP-digoxigenin nick end labeling (TUNEL) staining and immunohistochemistry.
Biochem/physiol Actions
Peroxidase (POD) helps in the dehydrogenation of several organic compounds like phenols, aromatic amines, hydroquinones etc.
Quality
Isoenzymes: >90% isoenzyme C (HPLC)
Purity number: 3.0 - 3.5 (A403/A275)
Preparation Note
Stabilizers: Product is stabilized with sucrose.
Working concentration: 1:4,000 to 1:10,000
For ELISA
Working solution: Preparation of the solutions, (15 to 25 °C)
1. 1 M Sodium carbonate/-hydrogencarbonate solution, pH 9.4
1 M Na2CO3: Dissolve 10.6 g Na2CO3 in 80 ml double-dist. water and make up to 100 ml.
1 M NaHCO3: Dissolve 8.4 g NaHCO3 in 80 ml double-dist. water and make up to 100 ml.
Adjust the pH of the NaHCO3 solution to 9.4 by adding Na2CO3 solution.
2. 100 mM Sodium carbonate/-hydrogencarbonate solution, pH 9.8
Dilute 10 ml solution 1 to 100 ml with double-dist. water.
3. 200 mM Sodium borohydride solution
NB: Prepare the solution immediately prior to use and keep cold on ice. Dissolve 8 mg NaBH4 in 1 ml cold double-dist. water.
4. 2 M Triethanolamine solution, pH 8.0
Dilute 2.66 ml triethanolamine with 3 ml double-dist. water, adjust the pH to 8.0 with 25% HCI and make up to 10 ml with double-dist. water.
5. 1 M Glycine solution, pH 7.0
Dissolve 0.75 g glycine in ca. 6 ml double-dist. water, adjust to pH 7.0 with 0.1 M NaOH, and make up to 10 ml with double-dist. water.
6. PBS (phosphate-buffered saline); glycine; pH 7.4.
10 mM Potassium phosphate, 200 mM NaCI, 10 mM glycine, pH 7.5.
- Solution A (K2HPO4): Dissolve 4.56 g K2HPO4 x 3 H2O, 23.4 g NaCI, and 1.5 g glycine in ca. 1,500 ml double-dist. water and make up to 2000 ml with double-dist. water.
- Solution B (KH2PO4): Dissolve 2.72 g KH2PO4, 23.4 g NaCI, and 1.5 g glycine in ca. 1,500 ml double-dist. water and make up to 2,000 ml with double-dist. water.
- PBS: Whilst controlling pH, add sufficient solution B to solution A until the pH is 7.4.
7. Antibody solution
0.3 ml required for each labeling reaction. The IgG concentration of the solution to be used is c = 4 mg/ml (3.8–42 mg/ml). This value is critical for the coupling and hence should be checked photo-metrically for every test and adjusted if necessary: A280nm, 1cm, 1 mg/ml = 1.40.
NB: Do not use preservatives (e.g., sodium azide) and stabilizers (e.g., albumin).
- Immunoglobulin, salt-free, lyophilized:
- Immunoglobulin in buffer:
Stability of the solutions
Solutions 1, 2, 4, 5 and 6 are stable for 1 week at 2 to 8 °C. Solutions 3 and 7 should always be prepared immediately prior to use.
Storage conditions (working solution): The reconstituted solution is stable for 3 months at 2 to 8 °C. The solution can be aliquoted and shock-frozen at -60 °C or below, and then stored at -15 to -25 °C; a loss of activity of 10–20 % can result.
Reconstitution
Dissolving the lyophilizate in 0.5 ml double-dist. Water produces a peroxidase concentration of 16 mg/ml.
Quality Level | 100, |
form | lyophilized |
specific activity | 800 units/mg protein (at 25 °C with ABTS and H2O2 as substrate (pH 5.0).) |
packaging | pkg of 8 mg (~40 mg lyophilizate) |
Торговая марка | Roche |
optimum pH | 6.0-6.5 |
shipped in | wet ice |
storage temp. | 2-8°C |
pictograms | GHS08 |
signalword | Danger |
hcodes | H317 - H334 |
pcodes | P261 - P280 - P284 - P304 + P340 - P333 + P313 - P342 + P311 |
RIDADR | NONH for all modes of transport |
WGK Germany | WGK 1 |
Flash Point F | does not flash |
Flash Point C | does not flash |
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