How Bacteriological Examination of Gelatin is done procedure for total bacteria count TBC 

Principal : Gelatin is a product obtained by partial hydrolysis of collagen derived from skin, white connective tissue, bones of animals. It is suitable in air when dry but is subjected to microbial contamination when moist or is sol’n. Therefore a limit test for the presence of total no. of bacterial per Gm total bacterial count is specified in the pharmacopoeia.
Preparation of sample :- Employ aseptic conditions through our use preferably a powdered sample. If the gelatin is in sheets, flakes shreds grind the material under aseptic conditions through a sterile grinder into a sterile bag or other sterile container. After mixing through by weigh 5 gm of the powdered sample and place it in a sterile dilution bottle containing 95ml of sterilized water. After the gelatin is thoroughly welted, place the container in a water bath heated to between 400 and 450 when the content become uniformly heated, shake well until solution is complete.
Dilution:- Dilute 20ml of this freshly prepared 1 in 20 solution. with 80ml of sterilized water to make a 1 in 100 solution. By decimal dilution prepare 1 in 1000 and10,000 dilution, of the dissolved gelatin. It the gelatin is known to be of good quality, the 1 in 20, 1 in 100 dilutions will suffice. The additional weaker solution are to be made for gelatin samples known or thought to posses a high bacterial content. Shake each dilution vigorously at least 25 times before a second dilution is made from it or before a sample is removed for plating.
Plating for Total count:- Use sterile pipette graduated deliver 1ml and glass covered Petri dishes 10cm in dia and 15min in depth for plating.
Plate out in duplicate 1ml each of the 1 in 20, 1 in 100 and other dilution if necessary.
Platting should be done immediately after the dilutions are prepared place 1 ml of the dilution. In a sterile Petri dish, add to the Petri dish 10ml of liquefied nutrient agar at a temp of 400. Raise the cover of the Petri dish just enough for the introduction at the pipette or culture medium. Flame the lips of all and flasks, test tubes and other containers used in delivering the medium mix the contents of Petri dish thoroughly by tilting and rotating the dish. All plates are to be solidified as quickly as possible and after inverting all glass covered plates, incubate them for seventhly two hours at 370 counts by performance the plates having between 30 and 300 colonies. Enumerate the express results in terms of 2.5 diameter magnification, with a focal distance of 8.89cm.

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