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AgriseraECL SuperBright (100 ml)

125 € / 100 €

AS16 ECL-S-100 

PRODUCT INFORMATION IN PDF

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AS16 ECL-S-100

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product information
Background

AgriseraECL SuperBright for Western Blot detection is a high quality substrate for detection of horseradish peroxidase enzyme activity at a femtogram detection levels. It is a ready to use 2 component system with low background and superior signal to noise ratios and highest sensitivity.

This reagent has superior stability and 18 months shelf life. Mixed working stock is stable for several days at ambient temperatures or when stored at 4°C.

Host
Clonality
Clone
Purity
Format
Quantity 2 x 50 ml, two component ready to use solutions, enough for 50 midi blots (6.8 x 8.1 cm),  which is 2754 cm2
Reconstitution
Storage

Highly stable for 18 month at 2°C to 8°C. Store in the dark. Mixed working stock is stable is stable for several days at ambient temperatures or when stored at 4°C. Agrisera ECL SuperBright possesses exceptional lot to lot consistency.

Tested applications

Western blot (WB)

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Additional information

Extreme low femto level sensitivity. Low background and superior signal to noise ratios.

HS code for this product is: 3822.00.0002

User Instruction

  • Store reagents A and B in the darkness at 4-8°C.
  • Mix equal volumes of reagent A and B in a clean container and equlibrate to room temperature 30 minutes before use.
  • Wash membrane with PBS or TBS -  prior to ading substrate, to remove any background prior to substrate contact.
  • Optimal results are obtained up to 20 minutes after substrate contact.
application information
Recommended dilution
Expected | apparent MW
Confirmed reactivity
Predicted reactivity
Not reactive in
Additional information
Selected references

Application example

Western blot detection using Agrisera ECLSuperBright reagent


10 µg of total protein from Arabidopsis thaliana leaf (1), Hordeum vulgare leaf (2) were extracted with Protein Extraction Buffer PEB (AS08 300). Samples were diluted with 1X sample buffer (NuPAGE LDS sample buffer (Invitrogen) supplemented with 50 mM DTT and heat at 70°C for 5 min and keept on ice before loading. Protein samples were separated on 4-12% Bolt Plus gels,  LDS-PAGE and blotted for 70 minutes to PVDF using tank transfer. Blots were blocked immediately following transfer in 5% non-fat milk dissolved in 20 mM Tris, 137 mM sodium chloride pH 7.6 with 0.1% (v/v) Tween-20 (TBS-T) for 1h at room temperature with agitation. Blots were incubated in Agrisera anti-RbcL primary antibody (AS03 037) at a dilution of 1: 20 000. 
The antibody solution was decanted and the blot was rinsed briefly twice, and then washed 1x15 min and 3x5 min with TBS-T at room temperature with agitation. Blots were incubated in secondary antibody (goat anti-rabbit IgG horse radish peroxidase conjugated, recommended secondary antibody AS09 602, Agrisera) diluted to 1:20 000 following by washes and detection using Agrisera ECL SuperBright detection reagent. Images of the blots were obtained using a CCD imager (VersaDoc MP 4000) and Quantity One software (Bio-Rad). Exposure time was 60 seconds.

Note:
In the barley sample, a well characterised 44 kDa degradation product is observed (Kokobun et al. 2002).

Caution:
Do not add any protein or HRP enzyme to reagent solution.
Optional: wash your tube, microtiter plate or membrane with 0.2 M solution of Sodium Phosphate, Dibasic, in Deionized Water. (Initial pH may be approximately 9). Bring the pH down to 8.4 with the slow addition of a 0.2M solution of Sodium Phosphate, Monobasic, in water.