The hepatitis B surface antigen (HBsAg) is an important marker to diagnose the infection of the hepatitis B in humans. The demonstration of HBsAg in the liver cell has been possible through special stains of the paraffin embedded sections. However, the results are not consistent by different staining method.
This study was undertaken to compare two most useful staining methods for the detection of HBsAg and also to find out the optional condition of these stains.
Original orcein staining(Shikata et al) is done at the pH 1 2 during 4 hours in orcein solution. This, however, was a non-specific stain for HBsAg and a similar type of result could be obtained even by hematoxylin and eosin. Further more it was difficult to obtain good contrast between HBsAg positive cells and the adjacent liver cells.
The orcein staining has been modified by the following conditions
1 . pH 1 -- 2, 60 minutes in orcein solution.
2 . pH 2 -- 3, 120 minutes in orcein solution.
Above the conditions, demonstrated HBsAg positive cells consistently. However, still orcein has some weak point such as difficulty in recognition of HBsAg through density, not contrast of color, non stability of dye solution for good staining. On the other hand, the other method, Victoria blue staining, was tried and following results were obtained.
1 . HBsAg positive cells stained blue.
2 . The other liver cells, including the cytoplasm and nucleus, stained red.
The HBsAg could be clearly demonstrated by Victoria blue stain in blue color against red cytoplasm and nucleus. So one could reconize HBsAg easily even in necrotic or necrobiotic cells.
3 . This staining method showed a wide secure differentiation time from 3 minutes to 15 minutes as well as a wide secure staining time from 1 day to 1 week.
4 . The Victoria blue staining required the oxidazing and reducing solutions to prepare a working solution on the day in use to obtain clear demonstration of the liver cell structure.
HBsAg positive rates in liver tissues by Victoria blue staining were 41.2% in chronic persistent hepatitis, 28.0% in hepatocellular carcinoma, 11.5% in liver cirrhosis, 8.3% in acute viral hepatitis, 6.7% in chronic activehepatitis.
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