Medical

The Triple Sugar Iron (TSI) Test  To determine the ability of an organism to ferment glucose, lactose, and sucrose, and their ability to produce hydrogen sulfide. Principle The Triple Sugar Iron (TSI) test is a microbiological test named for its ability to test a microorganism’s ability to ferment sugars and to produce hydrogen sulfide. An agar slant of a special medium with multiple sugars constituting a pH-sensitive dye (phenol red), 1% lactose, 1% sucrose, 0.1% glucose, as well as sodium thiosulfate and ferrous sulfate or ferrous ammonium sulfate is used for carrying out the test.  . Due to the building of acid during fermentation, the pH falls. The acid base indicator Phenol red. is incorporated for detecting carbohydrate fermentation that is indicated by the change in color of the carbohydrate medium from orange red to yellow in the pres...

Blood Agar (BAP) Blood Agar (BA) are  enriched medium  used to culture those bacteria or microbes that do not grow easily. Such bacteria are called “fastidious” as they demand a special, enriched nutritional environment as compared to the routine bacteria.  Composition of Nutrient Agar 0.5% Peptone 0.3% beef extract/yeast extract 1.5% agar 0.5% NaCl Distilled water  Blood Agar is made from Nutrient Agar more: 5% Sheep Blood pH should be from 7.2 to 7.6 (7.4) Preparation of Blood Agar 1.     Suspend 28 g of nutrient agar powder in 1 litre of distilled water. 2.     Heat this mixture while stirring to fully dissolve all components. 3.     Autoclave the dissolved mixture at 121 degrees Celsius for 15 minutes. 4.     Once the nutrient agar has been autoclaved, allow it to cool but not solidify. 5.   ...

Mueller Hinton Agar (MHA)       The major use of Mueller Hinton Agar is for antimicrobial susceptibility testing. Composition of MHA Ingredients   Beef Extract                                  2.00 gm Acid Hydrolysate of Casein         17.50 gm Starch                                           1.50 gm Agar                                             17.00 gm Distilled Water                             1000 ml Final pH 7.3 ± 0.1 at 25ºC Why Mueller Hinton agar is used for antibiotic susceptibility tes...

   Gram stain reagents These stains can be ordered as a complete kit from VWR (#l5204-004) or can be reconstituted as follows:   Crystal violet crystal violet (90% dye content)                             20.0 g ethanol (95%)                                                          200 mL ammonium oxalate                                         ...

   Cytochrome oxidase   – this test determines the presence of cytochrome oxidase enzymes.        Oxidase reagent Tetramethyl-P-Phenylenediamine Dihydrochloride         1.0g d-H 2 O                                                                               100 mL The use of an iron-containing metal inoculation loop can lead to a false- positive reaction. Use only plastic or platinum loops for this test. 1.       Add an inoculum of pure 18 -24 hour old bacterial culture to the...

     Catalase  test – this test determines bacterial production of catalase enzymes.   1.       Place a drop of hydrogen peroxide (3% H 2 O 2 - reagent grade) on a microscope slide or in the concave surface of a hanging drop slide. 2.       With a sterile loop, collect a sample of 18-24 hour old pure bacterial culture.   3.       Place the loop in the hydrogen peroxide.   4.       If the test is positive, there will be immediate bubbling or foaming, and liberation of O 2 gas. 5.       Record results. Positive :   If the mixture produces bubbles or froth, the organism is said to be 'catalase-positive (   Staphylococcus  and  Micrococcus are catalase-positive. Other catalase-positive organisms  include  Listeria ,  Corynebacterium diphtheriae ,  Bur...