Bio-Rad Silver Stain Plus Kit User Manual

Silver Stain Plus

Catalog Number
#161-0449

™

For Technical Service, call your local Bio-Rad office or
in the U.S. call 1-800-424-6723

Introduction

Silver Stain Plus™ is a quick, simple system for detecting
proteins or nucleic acids in polyacrylamide and dried
agarose gels after electrophoresis. Proteins and nucleic
acids can be visualized in 1 hour with very little hands­on time by employing a carrier-complex silver staining
chemistry similar to that developed by Gottlieb and
Chavko for detecting DNA in agarose gels.1 Silver staining
is very sensitive. Silver Stain Plus is 30 to 50-fold more
sensitive than Coomassie Blue R-250 dye and will detect
nanogram quantities of protein and DNA. To ensure best
results, always use high-quality deionized distilled water
and extremely clean glassware. Always wear gloves when
using the kit components and when handling gels.

Components

The Silver Stain Plus kit includes the following components
in quantities sufficient to stain 40 mini gels
(8 x 10 cm) or 13 conventional gels (16 x 16–20 cm):

• Fixative enhancer concentrate

• Silver complex solution (contains NH4NO3 and AgNO3)

• Reduction moderator solution (contains tungstosilicic

acid)

• Image development reagent (contains formaldehyde)

• Development accelerator reagent (contains Na2CO3)

• Empty 1 L bottle for development accelerator

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Additional Materials Needed

In addition to the components in the Silver Stain Plus kit,
the following materials are required:

• Reagent-grade methanol and acetic acid

• Deionized, distilled, or high-purity water

• Water of 18-megohm-cm resistivity recommended

• Polytetrafluoroethylene (PTFE)-coated stir bars

• Glass or plastic staining trays

All steps in this procedure require gentle agitation of the
gel in solution. For best results, a shaker table is
recommended.

Warnings and Precautions

Read each label in the kit before beginning the procedure.

Wear gloves, safety glasses, and protective clothing while
preparing and working with all the solutions in this kit.

The image development reagent should be used only in
areas with good ventilation. Avoid breathing vapors. Avoid
contact with skin. In case of contact with eyes, flush with
copious amounts of water and contact a physician.

See Material Safety Data Sheets for additional
information.

Storage

All reagents except the development accelerator solution
can be stored at room temperature. The development
accelerator solution should be stored at 4°C but allowed
to reach ambient temperature before use. This may be
done by aliquoting the volume needed and either warming
in a water bath for 2–5 min or allowing to stand at room
temperature for approximately 30 min before use.

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Recommendations for Optimal
Staining

• To prevent silver deposits on staining trays and
inconsistent staining, all containers used for mixing
and staining should be scrupulously clean. Glass,
polyethylene, or polypropylene containers may be
cleaned with 50% (approx. 8 N) nitric acid after
cleaning with laboratory detergent. Rinse thoroughly
with high-quality deionized water

• Throughout the procedure the gel should always be
completely submerged in solution. Gels that float on
the surface of the solution will not stain consistently
and will show background discoloration

• Avoid staining gels in direct sunlight or at
temperatures above 25°C

• Never touch gels with metal objects or bare skin.
PVC or latex gloves rinsed with deionized water
should be used if the gel must be handled. Perform
gel manipulations with glass or polyethylene rods, if
possible

• Stopping point: The fixative step is a desirable
stopping point if there is not enough time to complete
the procedure

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Reagent Preparation

Development Accelerator Solution

1. Prepare once for use with the entire contents of the
kit.

2. Place 950 ml of deionized distilled water in a

1 L cylinder containing a PTFE-coated stir bar.

3. Begin stirring and slowly add the entire
50 g contents of the development accelerator reagent.
Stir until dissolved.

4. Add water to make 1 L. Pour the solution into the
provided empty 1 L bottle labeled development
accelerator solution.

5. Store at 4°C. Use at room temperature.

If the entire solution will not be used within 3 months,
prepare the development accelerator solution according to
your use rate.

To prepare half the solution, add 25 g development
accelerator reagent to 475 ml deionized distilled water,
add water to make 500 ml.

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