The Kleihauer Betke Test:

A peripheral blood smear of the mom’s blood is prepared and fixed in 80% ethanol. It is then treated with acid (McIlvaine’s buffer) and stained with eosin and hematoxylin. Fetal cells will remain in tact and stain bright pink because of the high HbF/fetal hemoglobin content which resists acid elution. Adult hemoglobin in maternal cells will elute out, leaving faintly staining “ghost cells”. The percental of fetal cells per 2000 cells is used to determine how much extra WinRho (in 300mg vials) is required:

Vials = %fetal cells x maternal blood volume (~5000mL)/30mL

Rounded to the nearest whole number + 1 extra. But that one extra is usually the shot they automatically gave her after she gave birth.

The Rosette Test / Screen for fetal-maternal bleeds

After the baby is born, we take their cord blood (which is thick and gross, by the way) and do a forward ABORh on them. If they are D positive or weak D positive and the mother is negative, the hospital will do an onsite rosette test:

A 3% cell suspension of the mother’s blood is incubated with chemically modified anti-D which will bind to any infant cells. Unbound antibody is washed away and indicator cells (O positive cells treated with ficin to increase reactivity) will bind with infant cells to make little microscopic rosettes/clumps which you can look for with an inverted mic:

All eligible mothers will receive another shot of WinRho regardless of the result because the clumps can be hard to see. However, positive screens indicate a fetal bleed over 30mL (which occurs in about 0.3% of pregnancies), in which case, the mother might need a larger dose, determined by a Kleihauer-Betke (in tomorrow’s post). The WinRho is administered within 72hours because once mom’s discharged, it’s really hard to hunt a lady who just gave birth down just to give her another needle!

Chlamydospores in Candida albicans, wet mount.

The Dalmau test was used to help ID C. albicans once upon a time but phased out before I got into the labs. It was essentially growing a yeast in unfavourable environments to see if they formed chlamydospores in response to the stress. Cornmeal agar (minimal nutrient qualities) was used and the inoculum would be covered with a coverslip to decrease the amount of oxygen available to the growing yeast.