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Common Buffer Stock Solutions

For DNA gels

TBE 10X

Compound Mass Concentration
Tris base 121.1 g 1 M
Boric acid 61.8 g 1 M
EDTA (tetracid) 6.5 g 0.02 M

Adjust to 1 liter with nano-pure water. For electrophoresis, you can use both 1X or 0.5X.

TAE 50X

Compound Mass Concentration
Tris base 242 g 2 M
Glacial Acetic Acid 57.1 mL 1 M
EDTA (tetraacid) 16.3 g 0.02 M

Adjust to 1 liter with nano-pure water. For electrophoresis, use 1X.

denaturing PAGE gel stock solutions

Compound X% 6% 8% 15%
40% Acrylamide Bis 19:1 1.25*X mL 7.5 mL 10 mL 18.75 mL
10X TBE 5 mL
Urea 21 g

Adjust to 50 mL with nano-pure water.

For casting gels, use the volumes noted in the following table:

gel type spacers Volume needed
mini 1 mm 6 mL
1.5 mm 10 mL
midi 1 mm 27 mL
1.5 mm 40 mL

Then add 1/1000th of 25% APS and TEMED (1 uL per mL) and quickly cast the gel.

TLC stains

  • p-Anisaldehyde: This clear tan stain is prepared by dissolving 0.7 mL of p-anisaldehyde in 250 mL of EtOH containing 9.5 mL of conc. H2SO4 and 2.7 mL of acetic acid.
    This stain chars lavender and is good for oxygenated compounds.

  • Cobalt(II) chloride: This pink stain is prepared by dissolving ~2 g of CoCl2 in 100 mL of water and 10 mL of conc. H2SO4. The stain is used in conjunction with PMA.
    In cases where PMA is not effective enough, dipping the plate (already processed with PMA) in CoCl2 assists the staining process.
    The spots are often different colors against a pink or colorless background.

  • 2,4-Dinitrophenylhydrazine: This stain is prepared by dissolving 12 g of 2,4-dinitrophenylhydrazine in 200 mL of EtOH, 60 mL of conc. H2SO4 and 80 mL of water. The stain is effective for carbonyl containing compounds.

  • Ehrlich's Reagent: This stain is prepared by dissolving 1.0 g of p-dimethylbenzaldehyde in 75 mL of MeOH and 50 mL of concentrated HCl. The stain is effective for indoles, amines, ergot alcohols and pyrrolizidine alkaloids.

  • Ceric Ammonium Molybdate (CAM): This stain is prepared by dissolving 2.5 g of (NH4) 6-Mo7O24 and 1.0 g Ce(SO4)2 in 90 mL of water and 10 mL of conc. H2SO4. The plate is developed by heating on a hot plate.
    This stain chars blue and is good for polyhydroxylated compounds.

  • I2: A jar of I2 crystals or a jar of fine grade silica gel impregnated with iodine works well. The plate is simply allowed to stand in the jar of iodine vapor or is immersed in the silica gel for a few seconds. After examining the plate, the stain can be removed by heating the plate briefly on a hot plate. The plate can then be stained by other means.

  • Ninhydrin: This stain is prepared by dissolving 0.3 g of ninhydrin in 100 mL of n-BuOH or EtOH containing 1-3 mL of acetic acid. The plate is developed by heating on a hot plate and spots are shades of purple or brown. The stain is specific for amino acids, amino sugars and primary, secondary, and tertiary amines.

  • Phosphomolybdic acid (PMA): This green-yellow stain is prepared by dissolving ~3-4 g of PMA in 100 mL of EtOH. The plate is developed by heating on a hot plate. PMA is the most commonly used stain for organic materials, since it reacts with a wide variety of functional moieties. The spots are usually black against a yellow-green background. PMA is also effective with extremely dilute reaction samples and in this regard can be used semi-quantitatively.

  • Potassium Permanganate: This stain is prepared by dissolving 3 g of KMnO4 and 20 g of K2CO3 in 5 mL of 5% NaOH and 300 mL of water.

  • Vanillin: This light tan stain is prepared by dissolving 1-2 g of vanillin in 100 mL of EtOH containing 1 mL of conc. H2SO4. The plate is developed by heating on a hot plate. This stain is particularly good for hydroxyl and carbonyl containing molecules. The spots are often brightly colored. This may be an advantage when two molecules have the same Rf value.

Cooling Baths

Cooling baths can easily be created by combining a coolant (such as CO2 or N2) with a solvent that has a melting point at the appropriate temperature.

Further links

Chemistry

Biology

  • OpenWetWare, a site containing relevant buffer recipes and other neat information.