The various staining procedures that utilize silver are based on very different principles. Five categories of silver stains can be defined by the physicochemical procedures involved. These are argentaffin methods, argyrophil methods, impregnation stains, silver oxidation-reduction stains, and metallic-metallic interactions (autometallographic). In ASRBC laboratory Silver oxidation- reduction staining process is being used for DNA based hybrid purity test of some vegetables. Silver staining is a very sensitive method for detecting small amounts of proteins and low-molecular-weight nucleic acids in polyacrylamide gels. In general, the method is about 100 times more sensitive than Comassie blue for many proteins and several times more sensitive than EtBr staining for the visualization of polynucleotides. Besides, this staining process is very time sensitive and time has to be maintained to the exact. It has the advantages of allowing visualization of the sample without any specialized equipment and rendering gel backgrounds that are virtually colorless.
After completing PCR product’s polyacrylamide gel electrophoresis, gels have to be washed with milliq water and then the gel is dipped into acetic acid with gentle shaking for fixation. Next step is to add impregnate solution that contains silver nitrate (AgNO3). In this step, silver attaches with the phosphate group of DNA and form an insoluble silver phosphate (AgPO4). Then after rinsing with developer solution where sodium thyosulphate (Na2S2.5H2O) is present that perform oxidation-reduction reaction and stains the gel where DNA bands are present. Lastly again acetic acid is added to fix the stained gel.
Assistance of Tasnin Khan Eusufzai & Aqief Afzal, Research Assistants of ASRBC, ACI Ltd in preparing the article is highly appreciated.
Professor Lutfur Rahman
