Help for Chocolate agar plate

Derek Law dlaw at
Mon Nov 27 10:57:57 EST 2000

Chocolate agar is difficult to make and batch to batch variation is very common due to differences in temperature of the medium when blood is added, standing time after adding blood and the way the medium is mixed.

It is possible that the agar is being heated too much during stages 6 and 7, a 2 litre volume of agar takes time to heat up and cool down, potentially exposing the blood to overheating although the temperature used is relatively low. Try making the media in 4 x 500ml volumes and see what happens. If the media looks better this way it is due to overheating in a large volume.

Derek Law
Company Microbiologist


  Simon Lee <pedlcymd at> wrote in message news:8vt87u$fm5$1 at
  I want to solve the problem about the chocolate agar palte.
  The our method of preparation is belows.
  1. 2L ditilled water + 144 gm GC agar powder 
  2. dissolve fully with heating till boiling
  3. autoclave 
  4. cooling till 45 C degree
  5. add 5% defibrinated sheep blood
  6. heating to 65 C degree
  7. cooling till 45 C degree
  8. add IsoVitale X 1 vial
  9. pouring into plate.

  The problem is two
  One ; there are many sand like particle on the surface and inner side of agar plate. - I have heard that the problem is stirring during phase 5,6 ? Is it right ?
  Two ; the color of chocolate agar palte is too dark

  Please help me ASAP.
  Simon Lee

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