Small, wet, or off-color crystals may be cleaned by immersing them in distilled water for a short time, followed by drying with tissue or a paper towel. Add the sulfide to the oxygen-free water and swirl to dissolve. Dispense, seal, and autoclave. Always write the date on the labels, and discard the solution within 4 weeks. Remember that dispensing medium and sealing culture vessels are the most important steps for exclusion of Oxygen because these are the steps at which gaseous air is excluded from the culture vessels.
Air is a much more serious potential source of oxygen than is oxygen dissolved in liquid.
A given volume of air contains 30 to 40 times as much oxygen as does an equal volume of air-saturated water. To seal culture vessels, slide out the needle delivering oxygen-free gas and immediately insert the stopper. Do not press down on the stopper while removing the needle, but rather insert the stopper quickly and gently the instant the needle cannula has cleared. During autoclaving, dissolved CO 2 species leave solution into the gas phase, making the medium alkaline, causing precipitation of divalent cations. When the agar is cooled to 45 C and mixed, the bicarbonate buffer is re-established and these precipitated salts redissolve.