Sampling tools Sampling bags Solid and liquid food
Sampling tools �Sampling bags �Solid and liquid food or environmental samples �Sampling tube �Liquid samples �Sponge or cotton swab �For surface �Each group choose your own samples and collection tools. Food samples is preferred.
�Food samples �ready-to-eat food �It is better to store at room temperature for at least half day �Solid (e. g. sandwich, egg roll) �Liquid (e. g. soy milk, milk tea)
total plate count (TPC) aerobic plate count (APC) total mesophilic plate count �Total bacterial number is an important indication for food quality �Basic principle for TPC �Sampling size, 25 or 50 g �Solid sample �Add 9 times volume of buffer (25 g + 225 m. L wt: vol; 25 m. L + 225 m. L vol: vol) �Homogenizer or stomacher
Materials and methods �Food samples �store at room temperature �Depend on the food samples to decide the dilution range �Dilution range is from -1 to -4
Detection limit • No dilution – 1 CFU/ml; 10 CFU/g • -1 dilution – 10 CFU/ml; 100 CFU/g • However, we need 25 to 250 CFU in one plate • Thus, reliable detection limit is: • No dilution – 25 CFU/ml; 250 CFU/g • -1 dilution – 250 CFU/ml; 2500 CFU/g
procedures 1. Homogenize sample (25 g + 225 m. L phosphate buffer, 1: 9 dilution) 1. Homogenize for 2 min 2. For milk, no homogenization 2. Decimally serial dilution 3. Place one m. L of the undiluted homogenate into a Petridish (0 dilution) 1. Add agar to mix the sample: 混稀 pour plate 4. Place 0. 1 m. L of the undiluted homogenate onto a Petridish (-1 dilution) 塗抹: spread plate 5. Place 0. 1 m. L of -1 dilution the homogenate onto a Petridish (-2 dilution)
• Incubate the plates at 37 C for 24 (spread) to 48 (pour) hours • Count the colony (25 -250 colony/plate) • Calculate bacterial number in food sample – Solid food: average CFU number x dilution x 10 – E. g. 56 x 103 x 10 = 560000 = 5. 6 x 105 CFU/g = 5. 75 log 10 CFU/g – Milk: average CFU number x dilution – E. g. 56 x 103 = 56000 = 5. 6 x 104 CFU/g = 4. 75 log 10 CFU/m. L
Materials and methods �Food sample: �Each group decide their own sample. Usually ready-to-eat food has not high amount of bacteria. Thus, buy them one day before and place at room temperature. �Don’t buy cookies, bread, any food item with package. �One sample bag
�Dilution Buffer (PBS) 225 m. L �Dilution tubes �Plate (TSA or PCA)-6 plates (spread) �The dilution will be decided on the class depending on the food item. �Stomacher �Test tube racks
procedures �Purchase the food item and place into the sampling bag. �Record the time, place, food name, who sample it. �One the class, take 25 g from the food item. Do NOT just take one site, take several sites of the food item. �Add 225 m. L (1: 9), stomach the sample for at least 1 min.
�Dilution the samples according to the dilutions. �Spread plates. � incubate at 37 C for 24 hours. �Count the colony number at the suitable dilution (25 -250 CFU). �Calculate the bacterial counts in the sample.
Preparation of media(um) �Two types of agar �Tryptic Soy Agar (TSA) �Plate Count Agar (PCA) �Add agar powder into a bottle or flask according to the instruction on the bottle �TSA -40 g/L �PCA 23. 5 g/L
PCA �Pancreatic Digest of Casein. . . . 5. 0 g Yeast Extract ………. . . . 2. 5 g Dextrose. . . 1. 0 g �Agar. . . . 15. 0 g
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