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Antibodies

Aldolase Antibody Peroxidase Conjugated

Product Sizes
25 ul
200-1341S-25UL
About this Product
SKU:
200-1341S
Additional Names:
ALDOA|goat anti-Aldolase Antibody, HRP conjugated goat anti-Aldolase Antibody, Fructose-bisphosphate aldolase A, Muscle-type aldolase, peroxidase Aldolase
Application:
ELISA, Immunoprecipitation, Western Blot
Buffer:
0.02 M Potassium Phosphate, 0.15 M Sodium Chloride
CE/IVD:
RUO
Clonality:
Polyclonal
Conjugate:
HRP
Concentration:
1 mg/ml
Extra Details:
Aldolase is an enzyme that performs an aldol reaction (creating an aldol) or its reverse (cleaving an aldol). Aldolase plays a key role in glycolysis and gluconeogenesis. In addition, may also function as scaffolding protein. Belongs to the class I fructose-bisphosphate aldolase family.
Formulation:
0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2
Host:
Goat
Immunogen:
Aldolase [Rabbit Muscle]
Isotype:
IgG
Physical State:
Liquid
Reactivities:
Human, Rabbit
Shipping Conditions:
Dry Ice
Specificity:
Aldolase is an IgG fraction antibody purified from monospecific antiserum by a multi-step process which includes delipidation, salt fractionation and ion exchange chromatography followed by extensive dialysis against the buffer stated above. Assay by immunoelectrophoresis resulted in a single precipitin arc against anti-Peroxidase, anti-Goat Serum as well as purified and partially purified Aldolas
Storage Conditions:
-20[o]C diluted. Avoid freeze/thaw cycles.
Supplier:
Rockland Inc
Type:
Antibody: Polyclonal Antibody
IgG purified antibody to rabbit muscle aldolase (100-1141, 200-1141 and 200-1341) was used at a 1:1000 dilution to detect human aldolase by western blot. A whole cell lysate prepared from human derived A293 cells was loaded on a 4-12% tris glycine gradient gel for SDS-PAGE. The gel was transferred to nitro-cellulose using standard techniques. Antibody reaction with the membrane occurred overnight at 4° C in TTBS supplemented with 2% non-fat dry milk. Color was allowed to develop using SuperSignal West Pico Chemiluminescent Substrate (PIERCE). Other detection methods will yield similar results. This antibody clearly detects a band at ~41 kDa consistent with human aldolase.
Anti aldolase antibody - Immunoprecipitation and Western Blot. 300 ul aliquots of whole anti-aldolase antiserum (100-1141) were used to precipitate varying amounts of purified aldolase and precipitates with controls were compared by SDS-PAGE and Western blot. Samples shown in the image are: 1. Purified aldolase 2. 300 ul antiserum with no antigen (negative control) 3. 300 ul antiserum with ~100 ul aldolase (2.5 mg/ml) 4. 300 ul antiserum with ~200 ul aldolase (2.5 mg/ml) For the precipitation, 300 ul of antiserum and an equal volume of aldolase antigen in PBS was incubated ~24 hrs at 4°C, centrifuged for 6 minutes at 13K RPM, washed once with PBS, centrifuged and dissolved in 60 ul 0.1 N NaOH. 90 ul of PBS was added, the sample was divided in 2 portions, and an equal volume of reducing (+4% BME) or non-reducing 2X sample buffer was added. The reduced samples were boiled for five minutes, and all samples were run at 140 V for ~45 minutes on a 4-20% tris/glycine gradient gel. Gel was stained, destained and imaged (see attached) using standard protocols. Precipitation of aldolase was confirmed by comparison of increasing amounts of antigen with the control protein by SDS PAGE and observation of a 40-45 kD MW band corresponding to Aldolase. Additional higher and lower molecular weight bands correspond to serum proteins.
Anti aldolase antibody- Immunoprecipitation- Immunoprecipitation was performed with 300 ul of anti aldolase antiserum and an equal volume of varied amounts (diluted from a stock solution of ~2.5 mg/ml) of purified aldolase in PBS. Antibody/Antigen mixture was incubated ~24 hrs at 4°C, centrifuged for 6 minutes at 13K RPM, washed once with PBS, centrifuged and dissolved in 60 ul 0.1 N NaOH. 90 ul of PBS was added, the sample was divided in 2 portions, and an equal volume of reducing (+4% BME) or non-reducing 2X sample buffer was added. The reduced samples were boiled for five minutes, and all samples were run at 140 V for ~45 minutes on a 4-20% tris/glycine gradient gel. Gel was stained, destained and imaged(see attached) using standard protocols. Precipitation of aldolase was confirmed by comparison of increasing amounts of antigen with the control protein by SDS PAGE and observation of a 40-45 kD MW band corresponding to Aldolase. Additional higher and lower molecular weight bands correspond to serum proteins.