Goat anti-Mouse IgG (H&L), HRP conjugated
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For long time storage after reconstitution, dilute the antibody solution with glycerol to a final concentration of 50% glycerol and store as liquid at -20°C, to prevent loss of enzymatic activity.
For example, if you have reconstituted 1 mg of antibody in 1.1 ml of sterile water add 1.1 ml of glycerol. Such solution will not freeze in -20°C. If you are using a 1:5000 dilution prior to diluting with glycerol, then you would need to use a 1:2500 dilution after adding glycerol. Prepare working dilution prior to use and then discard. Be sure to mix well but without foaming.
500 femtomoles of His-tagged proteins IsiA, NifH, PsbA, PsbB and PetC were loaded per gel well in Agrisera PEB extraction buffer. Proteins were separated on 4-12 % NuPAGE PAGE Bis-Tris polycacrylamide gel (Invitrogen) and blotted 1h to PVDF. Blots were blocked with for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 1 000 for 1h at RT with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed once for 15 min and 3 times for 5 min in TBS-T at RT with agitation. Blot was incubated in secondary antibody (goat, anti-mouse IgG horse radish peroxidase conjugated, from Agrisera AS11 1772) diluted to 1:25 000 in 2 % ECL Advance blocking reagent for 1h at RT with agitation. The blot was washed as above and developed for 5 min with ECL according to the manufacturers instructions. Exposure time was 5 seconds.
Apparent molecular weight of recombinant proteins: IsiA - 27 kda, NifH - 34 kDa, PsbA - 30-37 kDa, PsbB - 40 kDa, PetC - 23 kDa.
Purity of this preparation is > 95% based on SDS-PAGE. Antibody concentration is 1.0 mg/ml. Antibody is supplied in 10 mM sodium phosphate, 0.15 M sodium chloride, pH 7.2.1 % (w/v) B, Protease/IgG free. Contains 0.1 % (v/v) Kathon CG as preservative of bacterial growth.
The antibody will detect all isotypes of mouse IgG.
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