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7490-OP OmniPur® Silver Pro Stain Kit - Calbiochem

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7490-OP
  
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      Aperçu

      Replacement Information
      Description
      Overview

      This product has been discontinued.





      Specifications:
      (Developer Solution) pH at 25°C: 10.00 min





      Intended for laboratory and manufacturing use only. Not for drug, food, or household use.
      Catalogue Number7490-OP
      Brand Family Calbiochem®
      References
      Product Information
      Quality LevelMQ100
      Applications
      Biological Information
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Storage and Shipping Information
      Storage +2°C to +8°C
      Do not freeze Ok to freeze
      Packaging Information
      Transport Information
      Supplemental Information
      Specifications
      Global Trade Item Number
      Référence GTIN
      7490-OP 0

      Documentation

      OmniPur® Silver Pro Stain Kit - Calbiochem Certificats d'analyse

      TitreNuméro de lot
      7490-OP
      Fiche technique

      Note that this data sheet is not lot-specific and is representative of the current specifications for this product. Please consult the vial label and the certificate of analysis for information on specific lots. Also note that shipping conditions may differ from storage conditions.

      Revision24-May-2011 JSW
      DescriptionOmniPur® Silver Pro Stain Kit is an extremely sensitive colorimetric staining procedure for the detection of subnanogram amounts of proteins resolved on polyacrylamide gels. With high sensitivity and very low background, it is ideal for the visualization of protein bands in dilute samples or for the detection of proteins present in trace amounts. It is generally about 100-fold more sensitive than the commonly used Coomassie® Blue stain1.

      The 4 hour staining procedure begins with a fixation step to reduce band diffusion and remove interfering substances. A sensitization process, which enhances binding of silver ions to proteins, follows fixation. The staining step allows silver ions to permeate the gel and bind preferentially to sulfhydryl and carboxyl side chains. In the final development step, silver ions are reduced to metallic silver to form precipitates that are visible as brown bands within the gel.

      Staining intensity varies by the type of protein in the sample. Acidic proteins and proteins with highly negatively charged sulfated sugar residues such as proteoglycans and mucins are not readily detected by silver stains.
      Recommended reaction conditions

      Materials supplied:
      • 10X Sensitizer Solution, 1 bottle, 250 ml
      • 10X Silver Stain Solution, 1 bottle, 250 ml
      • 5X Developer Solution, 2 bottles, 250 ml each

      Additional Materials Required but Not Supplied:
      • Methanol
      • Ethanol
      • 37% Formaldehyde
      • Glacial acetic acid
      • Distilled or deionized water

      Precautions and Recommendations:
      • All incubations should be conducted with continuous agitation on a shaker.
      • Solutions should be freshly prepared on day of use.
      • All steps should be performed at room temperature.
      • Water must be deionized and distilled.
      • Staining should be carried out in clean glass containers with sufficient quantities of solution to immerse the gel and allow it to move freely during agitation. Use separate containers for each gel.
      • Powder-free gloves should be worn during the procedure. Do not touch gel plates, staining dishes or gels with bare hands as prints will be visible on the stained gels. Rinse gloves in water between each step.
      • If crystals form in the developer solution during storage, warm gently until they re-dissolve.

      Reagent Preparation
      • Fixative (100 ml): prepare Fixative as follows:
      • 50 ml Methanol
      • 12 ml Glacial Acetic Acid
      • 1.35 ml 37% Formaldehyde
      • 36.65 ml Distilled/Deionized Water

      • 35% Ethanol (300 ml): prepare 35% ethanol as follows:
      • 81 ml 95% Ethanol
      • 219 ml Distilled/Deionized Water

      • 1X Sensitizer Solution (100 ml): prepare 1X Sensitizer Solution as follows:
      • 10 ml 10X Sensitizer Solution
      • 90 ml Distilled/Deionized Water

      • 1X Stain Solution (100 ml): prepare 1X Stain Solution as follows:
      • 10 ml 10X Stain Solution
      • 90 ml Distilled/Deionized Water

      • 1X Developer Solution (100 ml): prepare 1X Developer Solution as follows:
      • 20 ml 5X Developer Solution
      • 80 ml Distilled/Deionized Water

      • Stop Solution (100 ml): prepare Stop Solution as follows:
      • 50 ml Methanol
      • 12 ml Glacial Acetic Acid
      • 38 ml Distilled/Deionized Water

      Detailed Protocol
      1. Fix gel in 100 ml of Fixative 2 hr to overnight.
      2. Wash 3 times, 20 min per wash, in 35% Ethanol.
      3. Incubate gel in 100 ml 1X Sensitizer Solution for 2 min.
      4. Wash 3 times, 5 min per wash, in distilled/deionized water.
      5. Incubate gel for 20 min in 1X Stain Solution.
      6. Wash gel 2 times, 1 min per wash, in distilled/deionized water.
      7. Incubate gel in 1X Developer Solution until bands become visible (approximately 10 min for full development). Bands should appear dark brown against a pale background.

      Note:
      • The rate of band development is temperature dependent.
      • If the gel is over-developed, artifacts are present, the background is too dark, or if the bands are over-stained, the gel can be de-stained and restained as desired.

      8. Incubate for 20 min in Stop Solution to prevent further color development.
      9. Store at 4°C in 1% Acetic Acid.
      10. Gels may be photographed on a bright white light box.

      Troubleshooting Guide

      A.Background is too dark:
      1. Residual acid in gel
      • Increase washing time after the fixation step.

      2. Poor quality acrylamide
      • Acrylamide quality can affect the background appearance of a silver-stained gel. Use ultrapure grade acrylamide.

      3. Poor quality water
      • Use deionized/distilled water in all solutions.

      B. Negative staining
      1. Excess protein in bands.
      • Reduce the amount of protein applied to gel.

      C. Streaking or yellow background
      1.Excess reducing agents such as 2-mercaptoethanol or DTT.
      • Reduce the amount of reducing agent in sample buffer.

      D. Artificial bands with apparent molecular weights between 50-70 kDa
      1. Excess amounts of reducing agents such as 2-mercaptoethanol or DTT.
      • Lower the amount of reducing agent in sample buffer.
      Storage +2°C to +8°C
      Do Not Freeze Ok to freeze