micro lab sheet # 1 - Huda 3awamleh

بسم الله الرحمن الرحيم

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We will talk today about two things ;

*** first : (precautions in microbiology lab)

*our lab is teaching lab not diagnostic as in hospital

*in the lab we should have a. central ventilation system ; in order not to open windows to aviod dust contamination (contamination means: transfer of bacteria or pathogens from sample to the environment or from environment to the sample)

b. uv light ; we should turn the uv light off when we are in the lab bcz its affect our DNA and the purpose of this light is sterilization for the air

* we use the detol as disinfectant or we use 70%ethanol (esberto)

*we make autoclavy at 121 celsius to our lab rubbish

C. bensin burner ;we get a sterile area from his flame and by cleaning the desk by a detol

d. loop ; tool which is used to transfer a portion from the sample to culture media for diagnosis

* example of the samples : urine sample , stool sample , csf , throat swab , ear swab , eye swab , vaginal swab for female , urethral swab , blood culture…..

***second : the practical , we have two exercises 1.wet mount preparation 2.gram stain

1.wet mount preparation : bacteria are either motile like proteus or immotile like staphylococci . we have atube inside it there is aliquid culture media (culture media : 2l 2awsat 2l 3'eda2yeh tosta5dam le tanmet al bacteria) inside this media are the essential nutrients as protein source , carbon source ,carbohydrate surce ,nitrogen source , minerals , water, and we put a mixture of proteus and staphylococci on the tube , on the slide we will take from the mixture by the loop and we will cover the slide by cover slip and we will see it by the microscope on 40X we will see the immotile bacteria staphylococci round in shape and the motile bacteria proteus rod in shape (both of bacteria still alive).

*the purpose from this experiment is to study the shpe and the motiliy of the bacteria

2.gram stain : the same tube we will take from it and we make gram stain ,we spread the mixture on the slide then we do smear then airdry then heat fixation to fix the smear on the slide and by heat the bacteria will die after the heat fixation we will add a blue stain called crystal violet for a minute then we washed by water then we add a gram iodine for a minute too the iodine will make acomplex with crystal violet in the gram + bacteria then we wash by tab water then we add 95%ethanol for decolorization the ethanol dissolve lipid and in this step the gram +will appear violet and the gram – will appear colorless we wash the slide then add safranin stain red in color to stain the gram – bacteria then wash then dry and examine by oil immersion lense 100X .. we will see the staphylococci round in shape, violet in color, in cluster, gram +,,, proteus rod,red,gram - .

*the purpose ;study the shape and size and arrangment of bacteria

*the bacteria either gram + or gram - , the bacteria that doesn’t accept gram stain we will use to stain it special stain.

*extra info. 1. We should sterilize the loop before and after usage untill its become red in color so its become sterile.

2.the circle of the loop is standared either 1/100 or 1/1000 or 1/500 ml from the sample . we usethe circle 1/100 ml .

3. we put vasleine on the border of the cover slip to prevent evapration before put it on the slide.

4.how to make crystal violet , we dissolved a specific quantity from a sepcific powder in ethanol and we make filtration.

5. we use diamond pen instead of marker to write on the slide.

6. the wet mount slide will appear shining in the microscope bcz of the refliction of the light (all bacteria are colorless).



Lab. Micro no.1

Huda awamleh.;)