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MORIMASA MATS UTA AND MAYUMI MATSUTA
- 20X SSC solution
- 175.3 g NaCl (MW 58.44)
- 88.2 g Na citrate (MW 294.1)
- Volume to 800 ml with distilled water.
- Adjust pH to 7.0 with 1M HCl.
- Adjust final volume to 1 liter.
- Autoclave if desired.
- PN buffer: 10 liter
- Make 10 liters 0.1 M Na2HP04 (dibasic). If using heptahydrate, MW 268.07: 268.07 gllO liters distilled water.
- Make 600 ml 0.1 M Na2HP04 (monobasic). If using monohydrate, MW 137.99: 8.28 g/600ml distilled water.
- Titrate the dibasic solution (pH>9) by adding the monobasic solution (pH -4.5) until a pH of 8.0 is reached. Keep
track of the volume of monobasic solution added; not all of
it will be used.
- Add Nonidet P-40 (NP-40) to 0.05% (should be approximately 5.25 ml). NP-40 does not influence the pH reading.
This buffer can be stored at room temperature indefinitely.
- PNM buffer:
- Add 5% nonfat dry milk to PN buffer + 0.02% NaAzide,
heat to 37°C, and leave on lab bench overnight, perhaps
longer. Spin in a tabletop centrifuge to pellet the solids.
Remove the supernant and store at 4°C or -20°e. If
more solids form, repeat spin.
- Antifade:
- 100 mg p-phenylenediamine dihydrochloride (Sigma
p1519) in 10 ml PBS
- Adjust to pH 8 with 0.5 M Carbonate-bicarbonate buffer
(0.42 g NaHC03 in 10 ml water, pH to 9 with NaOH).
- Add to 90 ml glycerol. Filter through 0.22 Ilm to remove undissolved particulates. Store in aliquot with the desired
counterstains in the dark at -20°e. This solution darkens
with time, but still remains effective. It can be used after more
than 6 months storage. 0. Immuno. Method 43,349,1981)
- Ethanol dehydrate solution
- For final concentrations of 70, 85 and 100%. Prepare v/v
dilutions of 100% ethanol with purified water. Use solution
for up to seven days and then discard. If solution evaporates
or becomes diluted, replace with fresh solution. Between
periods of use, store at room temperature.
MORIMASA MATS UTA AND MAYUMI MATSUTA
- 20X SSC solution
- 175.3 g NaCl (MW 58.44)
- 88.2 g Na citrate (MW 294.1)
- Volume to 800 ml with distilled water.
- Adjust pH to 7.0 with 1M HCl.
- Adjust final volume to 1 liter.
- Autoclave if desired.
- PN buffer: 10 liter
- Make 10 liters 0.1 M Na2HP04 (dibasic). If using heptahydrate, MW 268.07: 268.07 gllO liters distilled water.
- Make 600 ml 0.1 M Na2HP04 (monobasic). If using monohydrate, MW 137.99: 8.28 g/600ml distilled water.
- Titrate the dibasic solution (pH>9) by adding the monobasic solution (pH -4.5) until a pH of 8.0 is reached. Keep
track of the volume of monobasic solution added; not all of
it will be used.
- Add Nonidet P-40 (NP-40) to 0.05% (should be approximately 5.25 ml). NP-40 does not influence the pH reading.
This buffer can be stored at room temperature indefinitely.
- PNM buffer:
- Add 5% nonfat dry milk to PN buffer + 0.02% NaAzide,
heat to 37°C, and leave on lab bench overnight, perhaps
longer. Spin in a tabletop centrifuge to pellet the solids.
Remove the supernant and store at 4°C or -20°e. If
more solids form, repeat spin.
- Antifade:
- 100 mg p-phenylenediamine dihydrochloride (Sigma
p1519) in 10 ml PBS
- Adjust to pH 8 with 0.5 M Carbonate-bicarbonate buffer
(0.42 g NaHC03 in 10 ml water, pH to 9 with NaOH).
- Add to 90 ml glycerol. Filter through 0.22 Ilm to remove undissolved particulates. Store in aliquot with the desired
counterstains in the dark at -20°e. This solution darkens
with time, but still remains effective. It can be used after more
than 6 months storage. 0. Immuno. Method 43,349,1981)
- Ethanol dehydrate solution
- For final concentrations of 70, 85 and 100%. Prepare v/v
dilutions of 100% ethanol with purified water. Use solution
for up to seven days and then discard. If solution evaporates
or becomes diluted, replace with fresh solution. Between
periods of use, store at room temperature.