Plot No. C40, Road No. 21Y, MIDC, Wagle Industrial Estate,
Thane(W)-400604, MH, India.
Your enquiry has been submitted
Something problem with the server. Try again.
Intended Use
For alpha-toxin detection in Clostridium perfringens.
Composition**
| Ingredients | g/L |
|---|---|
| Sodium chloride | 8.500 |
| Agar | 15.000 |
| Final pH (at 25°C) | 7.0±0.2 |
**Formula adjusted, standardized to suit performance parameters
Directions
Suspend 23.5 grams in 1000 ml purified/distilled water. Heat to boiling to dissolve the medium completely. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. After cooling to 45-50°C, add blood to give final concentration of 5% v/v. Mix well and pour into sterile Petri plates.
Principle And Interpretation
A heat-labile enterotoxin produced only by sporulating cells (1) induces the major symptoms of diarrhea in perfringens poisoning. The enterotoxin appears to be released in vivo in the intestine by the sporulating organisms (2). Hence alpha toxin can be used as an index for detecting the presence of Clostridium perfringens in food (3). However, the viability of C. perfringens cells are lost if the suspected food samples are frozen (4). Saline Agar Base with blood is used to measure the haemolytic activity of alpha toxin (5,6,7).
Sodium chloride provides essential ions. Red blood cells are added in the medium to examine haemolytic reactions, which indirectly helps in detection of alpha toxin.
Additional Test
The plates containing 7 ml of medium is dried overnight at room temperature and stored at 4°C till use. Just prior to use, test wells are cut in the agar using a template space of test wells, 3 cm apart and 2 cm from the edge of the plate. Make 2 additional wells 3 cm apart near the centre of the plate. Peripheral wells of duplicate plates are filled with the undiluted extract (alpha toxin extraction) and eight twofold dilutions of extract. To determine whether the haemolysis caused by the extract is due to alpha toxin, a portion of the 1:2 dilution of the extract is mixed with C. perfringens alpha toxin and with C. perfringens type A diagnostic antiserum containing alpha toxin and placed in the two center wells. The plates are incubated for 24 hours at 35°C and examined for haemolytic zones surrounding the wells. A 1 mm zone of haemolysis is considered as significant (5).
Type of specimen
Clinical samples - faeces; Food and dairy samples.
Specimen Collection and Handling
For clinical samples follow appropriate techniques for handling specimens as per established guidelines (8,9).
For food and dairy samples, follow appropriate techniques for sample collection and processing as per guidelines (2,7,10,11). After use, contaminated materials must be sterilized by autoclaving before discarding.
Warning and Precautions
In Vitro diagnostic Use. For professional use only. 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 clinical specimens. Safety guidelines may be referred in individual safety data sheets.
Limitations
- Further biochemical and serological tests must be carried out for further identification.
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
White to light yellow homogeneous free flowing powder
Gelling
Firm, comparable with 1.5% Agar gel
Colour and Clarity of prepared medium
Basal Medium yields light yellow coloured, clear gel. On addition of red blood cells, red coloured opaque gel forms in Petri plates
Reaction
Reaction of 2.35% w/v aqueous solution at 25°C. pH: 7.0±0.2
pH
6.80-7.20
Cultural Response
Cultural characteristics observed after an incubation at 35-37°C for 18-24 hours with added red blood cells.
| Organism | Inoculum (CFU) | Haemolysis |
|---|---|---|
| Clostridium perfringens ATCC 12924 | 50-100 | positive reaction |
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 clinical sample must be decontaminated and disposed of in accordance with current laboratory techniques (8,9).
Reference
- śDuncan C. L., 1973, J. Bacteriol., 113:932.
- Salfinger Y., and Tortorello M.L., 2015, Compendium of Methods for the Microbiological Examination of Foods, 5th Ed., American Public Health Association, Washington, D.C.
- Harmon S. M., Kantler D. A., 1970, Method for Estimating the presence of Clostridium perferingens in Food.
- Hall H. E., 1968, J. Asst office Agar. Chem: 51: 1338-134.
- Dr. Williams Horwitz, (Ed.), 2000, Official Methods of Analysis of AOAC International, Maryland.
- Noyes N. E. and Easterling R., 1967, J. Bacteriol., 93:1254-1261.
- Sheldon D. R., Moskowitz M. and Daercerell M. W., 1958, J. Bacteriol., 77:375 - 382.
- Isenberg, H.D. Clinical Microbiology Procedures Handbook 2nd Edition.
- 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, 11th Edition. Vol. 1.
- American Public Health Association, Standard Methods for the Examination of Dairy Products, 1978, 14th Ed., Washington D.C.
- Wehr H. M. and Frank J. H., 2004, Standard Methods for the Microbiological Examination of Dairy Products, 17th Ed., APHA Inc., Washington, D.C.
| Product Name | Saline Agar |
|---|---|
| SKU | M942 |
| Product Type | Regular |
| Physical Form | Powder |
| Origin | Chemically defined |
| Packaging type | HDPE |
| References | 1. Duncan C. L., 1973, J. Bacteriol.,113:932. |
| Customized Product Available | No |
Write Your Own Review