TidyBlot WESTERN BLOT DETECTION REAGENT Accessory Reagent

TidyBlot WESTERN BLOT DETECTION REAGENT: HRP

Cat. Num. AAA50247

• Applications: Western Blot
• Synonyms: TidyBlot WESTERN BLOT DETECTION REAGENT; N/A; TidyBlot WESTERN BLOT DETECTION REAGENT: HRP; TidyBlot WESTERN BLOT DETECTION REAGENT accessory reagent

• Specificity: TidyBlot
• Form/Format: HRP
• Concentration: 40 mug/ml (varies by lot)

• Applicable Applications for TidyBlot WESTERN BLOT DETECTION REAGENT accessory reagent: WB (Western Blot)
• Instructions For Use: When using a 1/200 dilution (25ul in 5ml milk protein), TidyBlot Western Blot Detection Reagent: HRP is sufficient to perform approximately 20 mini Western Blots.; 1. Dilute primary antibody (see technical advice section) to appropriate concentration in blocking buffer.; 2. Incubate blot with primary antibody.; 3. Wash blot.; 4. Dilute TidyBlot Western Blot Detection Reagent: HRP For best results, optimize dilutions for each experiment.; 5. Incubate blot with TidyBlot Western Blot Detection Reagent:HRP.; 6. Wash blot.; 7. Prepare and add HRP substrate according to manufacturers instructions.; 8. Develop blot.
• Preparation and Storage: Store at +4 degree C. DO NOT FREEZE. This product should be stored undiluted. Should this product contain a precipitate we recommend microcentrifugation before use. ; Shelf Life: 12 months from date of despatch.

Application Data

(HeLa (lane 2) cell lysate was run under reducing conditions on SDS PAGE using the V3 system and transferred to a PVDF membrane. Precision Plus molecular weight marker was run in lane 1. Rabbit anti Human GAPDH was used as the primary antibody at a dilution of 1/1000 in lane 2. TidyBlot was used at 1/200 as the secondary detection reagent. Visualization was carried out using the ChemiDoc with automatic exposure)

Application Data

(25 ug of HeLa cell lysate spiked with 3.5 ug of mouse IgG was run in lanes 2, 3, 5 and 6 under reducing conditions on SDS PAGE using the V3 system and transferred to PVDF membrane. Precision Plus MW marker was run in lanes 1 and 4. Mouse anti actin gamma (VMA00049) was used to detect actin gamma (approximate MW of 46 kDa) in lanes 2 and 5 only. Lanes 3 and 6 were used as controls to detect the background signaling of the secondary detection agent (no primary antibody control).. TidyBlot Western Blot Detection Reagent:HRP was used as the secondary reagent in lanes 2 and 3 and an HRP conjugated anti-mouse IgG heavy and light chain antibody was used as the secondary in lanes 5 and 6. The Blot was visualized with the Chemidoc. The TidyBlot Western Blot Detection Reagent:HRP only bound to the native actin gamma antibody whereas the standard anti-Mouse IgG heavy and light recognized both the native and denatured antibodies)

Related Product Information for TidyBlot WESTERN BLOT DETECTION REAGENT accessory reagent

N.B. Reactivity of TidyBlot Western Blot Detection Reagent:HRP to mouse IgG1 can be variable. We recommend customers assess compatibility with their IgG1 antibodies via dot blot assay. TidyBlot Western Blot Detection Reagent:HRP specifically binds to native (non-reduced) antibodies. In contrast to conventional secondary antibodies, TidyBlot therefore enables the detection of immunoblotted target protein bands without interference from denatured IgG. Sources of denatured IgG include endogenous IgGs present in cell/tissue lysate and primary antibodies released from beads during immunoprecipitation experiments. The latter are particularly obstructive when detecting of (co-)immunoprecipitated proteins as IgG heavy (~50 kDa) and light chains (~25 kDa) may mask the protein(s) of interest.