Mast cell staining

From:"Timothy Martin" <>

Could anyone give me some information to help clear up and  
further understand a rather important bone of contention regarding  
staining properties and characteristics of the two different types of 
mast cells found in guinea-pig lungs and trachea. I have recently 
had  my PhD oral exam in Cardiff at the Welsh School of Pharmacy and 
the external examiner was took exception to the staining protocol 
(Find theprotocol I used  below) that I used to stain dispersed 
guinea-pig lung and racheal  mast cells  

it was first stated that the aclian blue and safranin should not be 
added at the same time but stained firstly with aclcian blue and then 
counterstained with safranin. Secondly it was claimed that guinea-pig 
connective tissue mast cells do not stain red with safranin O anyway, 
and thirdly it was claimed that fixing the cells was essential for 
the staining of these cells that without it the cells would not 
stain. He was therefore doubting my interpretation of my results 
which were that guinea-pig pig connective tissue mast cells stain red 
with safranine O whislt mucosal mast cells stain blue with  alcian 

Could you therefore give me any comments on the above, imparticular 
the importance of fixing cells on the staining properties of the 
alcian blue safranin combined stain on guinea-pig airway mast cells, 
and your opinion on whether you think the above protocol was suitable 
for the staining of guinea-pig mast cells. Also could you give me any 
further information regarding guinea-pig mast cells from the airways 

With kind regards  

Timothy Martin 
Department of Pharmacology 
Welsh School of Pharmacy 
Cathays Park 
CF10 3XF 


Guinea -pig lungs and trachea were first isolated and digested using 
collagenase and BSA. After filtering and spinning down the cells, 
were not fixed and  were stained in a solution which contained 0.36%
Alcian blue and 0.2% 
 Safranine O. This stain was made up in acetate buffer. 

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