Folic Acid Inoculum Medium

Intended Use

Recommended for preparation of inoculum of Enterococcus hirae ATCC 8043.

Composition

Final pH (at 25°C): 6.8±0.2

**Formula adjusted, standardized to suit performance parameters

# Equivalent to Peptonized milk

Directions

Suspend 38.0 grams in 1000 ml purified/distilled water. Heat if necessary to dissolve the medium completely. Distribute in tubes. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes.

Principle And Interpretation

An important part of any assay is the maintenance and inoculum preparation of the test organism. Folic Acid Inoculum Medium is used for the preparation of inoculum to be used in the assay of the vitamins. Folic Acid Inoculum Medium is formulated as described by Kavanagh (3) and recommended by AOAC (4) for inoculum preparation of Enterococcus hirae ATCC 8043, the test organism for Folic Acid Assay Medium (3).

Yeast extract and Peptonized SM powder supply mainly the nitrogenous nutrients, vitamins and minerals essential for the growth of the test organisms. Dextrose is the energy source in the medium while tomato juice provides the growth factors. Polysorbate 80 maintains the surface tension of the medium to the optimal level while phosphate serves as buffering to the medium.

Extreme care should be taken to avoid contamination of media or glassware used for the assay. Detergent free clean glassware should be used. Even small amount of contamination by foreign material can lead to erroneous results.

Type of specimen

Isolated Microorganism

Specimen Collection and Handling

Inoculate 10 ml of Folic Acid Inoculum Medium with an 18-24 hours old culture from Folic Acid Culture Agar (M134). Incubate at 35-37°C for 18-24 hours. Centrifuge the growth and resuspend the sediment in 10 ml of 0.85% sterile saline, after decanting the supernatant. Repeat washing with saline, two more times. Dilute 1 ml of the washed cell suspension with 99 ml of 0.85% sterile saline (1:100). Adjust the inoculum concentration as per requirement or standard reference (4).

Warning and Precautions

Read the label before opening the container. Wear protective gloves/protective clothing/eye protection/ face protection. Follow good microbiological lab practices while handling specimens and culture. Standard precautions as per established guidelines should be followed while handling specimens. Safety guidelines may be referred in individual safety data sheets.

Limitations

1. Extreme care should be taken to avoid contamination of media or glassware used for the assay.
2. Detergent free clean glassware should be used.

Performance and Evaluation

Performance of the medium is expected when used as per the direction on the label within the expiry period when stored at recommended temperature.

Quality Control

Appearance: Cream to yellow homogeneous free flowing powder

Colour and Clarity of prepared medium: Medium amber coloured, clear to slightly opalescent solution in tubes

Reaction: Reaction of 3.8% w/v aqueous solution at 25°C. pH: 6.8±0.2

pH: 6.60-7.00

Cultural Response: Cultural characteristics observed after an incubation at 35-37°C for 18-24 hours.

Key: (*) Corresponding WDCM numbers. (#) Formerly known as Lactobacillus casei

Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 20-30°C. Use before expiry date on the label. On opening, product should be properly stored dry, after tightly capping the bottle in order to prevent lump formation due to the hygroscopic nature of the product. Improper storage of the product may lead to lump formation. Store in dry ventilated area protected from extremes of temperature and sources of ignition. Seal the container tightly after use. Product performance is best if used within stated expiry period.

Disposal

User must ensure safe disposal by autoclaving and/or incineration of used or unusable preparations of this product. Follow established laboratory procedures in disposing of infectious materials and material that comes into contact with sample must be decontaminated and disposed of in accordance with current laboratory techniques (1,2).

Reference

1. Isenberg, H.D. Clinical Microbiology Procedures Handbook 2^nd Edition.
2. Jorgensen, J.H., Pfaller, M.A., Carroll, K.C., Funke, G., Landry, M.L., Richter, S.S and Warnock., D.W. (2015) Manual of Clinical Microbiology, 11^th Edition. Vol. 1.
3. Kavanagh F., 1963, Analytical Microbiology, Academic Press, New York.
4. Williams. (Ed.), 2005, Official Methods of Analysis of the Association of Official Analytical Chemists, 19^th ed., AOAC, Washington, D.C.