Why Is My Wright’s Or Wright-Giemsa Stain Too Blue?

Wright’s stain and Wright-Giemsa stain were created to make blood cell morphology more visible. They contain both eosin and methylene blue to better differentiate neutrophils, cytoplasm, and other cell parts within blood and bone marrow specimens. Consistent quality in these stains is essential. If the basophilic or eosinophilic staining is too strong, it can cause issues for pathologists that are accustomed to evaluating specimens with a different stain appearance. 

This post covers how to adjust staining protocols for specimens that are “too blue” or “too pink” after using a Wright’s stain or Wright-Giemsa stain.

Adjust basophilic staining that’s too strong or “too blue”

This issue is most commonly observed with classical stains conducted on an automated slide stainer. If you’re seeing nucleus staining that is “too blue”, we have some solutions you can try. 

1. Fix the specimen to correct degradation. If degradation of the specimen causes too much basophilic intensity due to a delay between fixation and staining, proceed by fixing a fresh smear and then move forward with your normal staining protocol. Adjust your procedure in the future to avoid this issue.
2. Reduce time in stain/buffer mix. You might naturally think to reduce time in the full 100% stain solution, but in reality, you should reduce time in the stain/buffer solution, since this is where the staining of the specimen actually occurs.
3. Try switching from 7.2 pH to 6.8 pH buffer. If you’re using a 7.2 pH buffer solution, switch to 6.8. Switching from a 7.2 pH buffer to 6.8 pH buffer will lower the pH of the buffer to be more eosinophilic (instead of basophilic/too blue). This will cause more of the intensity or distinction in the red blood cells and eosinophilic morphology rather than in the basophilic morphology.
4. Reduce the concentration of stain in the stain/buffer solution from 1:5 (20% stain, 80% buffer) to 1:10 (10% stain, 90% buffer). Ensure that the buffer/stain mix ratio has not been altered by over-use and to be safe, swap the solution for a new mix. This should be done every six hours to retain the buffer ratio.

For more suggestions on adjusting your staining techniques to meet your requirements download our eBook, Lab Technician’s Guide to Troubleshooting Common Issues with Biological Stains, call one of our application experts at 800-441-0366 or email info@ethosbiosciences.com. We love to help.