Inoculated agar surface

Paper disks are placed on the inoculated agar surface... 

Place six cylinders on the inoculated agar surface so that they are at approximately 60° intervals on a 2.8 cm. radius. Use three plates for each sample. FiU three cylinders on each plate with the 20 /ig. r milliliter standard and three cylinders with the 20 /ig. per milliliter (estimated) sample, alternating standard and sample. At the same time prepare a standard curve using concentrations of the standard of 36.0, 32.0, 28.0, 24.0, 20.0,16.0,12.0,10.0, and 8.0 />g. per milliliter in 1% phosphate buffer, pH 6.0. A total of 24 plates is used in the preparation of this standard curve, three plates for each solution, except the 20 /ig. per milliliter solution. The latter concentration is used as the reference point and is included on each plate. On each of three plates fill three cylinders with the 20 /ig. per milliliter standard and the other three cylinders with the concentration of the standard under test. Thus, there will be seventy-two 20 Mg. determinations and nine determinan tions for each of the other points on the curve. Incubate the plates for 16-18 hours at 32-35 °C. and measure the diameter of each zone of inhibition. Correct the average value for each point in the same manner as described for the penicillin assay (section 11,1). 

Place six cylinders on the inoculated agar surface so that they are at approximately 60° intervals on a 2.8-cm. radius. Use three plates for each sample. Fill three cylinders on each plate with the 100 units per milliliter standard and three cylinders with the 100 units per milliliter (estimated) sample, alternating standard and sample. A standard curve is prepared in the same manner as described for penicillin (Section 11,1), using standard concentrations of 180, 160, 140, 120, 100, 80, 60, 40, and 20 units per milliliter in 1% phosphate buffer, pH 6.0. Similarly, following the procedure of the penicillin assay, sample potencies are then calculated. 

Inoculate the surface of tryptone soya agar slant for bacteria and Sabouraud dextrose agar slant for fungi from recently revived stock culture of each of the test microorganisms. 

Plastic test discs are placed on the mineral salt agar surface and spray inoculated with a mixed fungal spore suspension. The plates are incubated at 20-25 C for four weeks or more. The degree of growth on the plastic and diameter of any zone of inhibition are recorded. 

Figure 1. Callus conditioning method. A a piece of callus, of about 100 mg fresh weight, is placed into a glass ring (22 mm diameter) previously sealed on the bottom by dialysis membrane. B the ring is placed on the surface of the inoculated agar medium.

To determine the antibacterial activity of the treated textile fabrics AATCC Test Method 147 is commonly used. In this method, an agar surface is inoculated by making a parallel streak. The sample is then pressed onto the plate, which is inoculated and incubated at 37 °C and the antibacterial activity is estimated by observing the decrease in growth of the organism from one end of each streak to the other end and from one streak to the next streak and by determining the size of the zone of inhibition. 

Inoculate a CSA plate from a thawed TSB stock of S. aureus using a sterile swab. Cover the agar surface completely and allow any liquid to absorb into the agar. 

Inoculate an individual TSA + 7.5% NaCl (see Note 25) bacterial agar plate with 100 pL (see Note 19) of supernatant from each sample and spread the solution evenly on the agar surface with a disposable spreader. 

With sterile forceps, carefully place sterile coverslip over inoculated agar square taking care not to leave exposed agar surface. 

Using a sterile loop, transfer a yeast colony onto the agar cube. With sterile forceps, carefully place a sterile cover slip over inoculated agar cube, taking care not to leave an exposed agar surface. 

Assay Technic. Add 20 ml. of base agar to each petri dish (20 X 100 mm.). Distribute the agar evenly in the plates and allow it to harden. Add 0.5-1.0 ml. of the Sarcina lutea suspension to each 100 ml. of agar (melted and cooled to 48°C.). Add 4.0 ml. of this seed agar to each plate, tilting the plates back and forth to spread the inoculated agar evenly over the surface. 

This material was made up with distilled water to provide 41 g per liter, and the mixture was adjusted to pH 7.0 with potassium hydroxide solution. To the mixture were added per liter 5.0 g of calcium carbonate and 7.5 ml of soybean oil. 2,000 ml portions of this medium were then added to fermentation vessels, equipped with stirrers and aeration spargers, and sterilized at 121°C for 60 minutes. After cooling the flasks were inoculated with a suspension of strain No. ATCC 11924 of Streptomyces lavendulae, obtained from the surface of agar slants. The flasks were stirred for 4 days at 28°C at approximately 1,700 rpm. At the end of this period the broth was found to contain cycloserine in the amount of about 250 C.D.U./ml of broth. The mycelium was separated from the broth by filtration. The broth had a pH of about 7.5. Tests showed it to be highly active against a variety of microorganisms. 

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