Babesia divergens was cultivated in RPMI 1640
(25 mM HEPES) supplemented with 10% human serum (RPMI-10%
HS) with a high percentage of parasitized erythrocytes (PPE)
([ges ]40%). Standardization of in vitro tests,
purification of
exoantigens, biochemical studies and the safety of the culture handler
motivated the development of a serum-free defined
medium. Removal of serum greatly reduced the PPE but, after a period of
adaptation, the culture was continuous and the
parasite was able to develop a 3% routine PPE. Addition of vitamins or
reduced
glutathione in basal medium (RPMI)
did not improve the PPE. The supplementation of basal medium with lipidic
carrier (Albumax I or bovine serum
albumin–Cohn's fraction V) promoted the growth of B.
divergens with high PPE (>30%) close to those obtained in
RPMI–10% HS. Neither protein nor lipid fractions alone were able
to
restore the growth of B. divergens. Nevertheless,
the whole lipid fraction from serum or Albumax I added to delipidated albumin
partially restored the growth (7% PPE),
indicating that the presentation of specific lipids by a carrier is
crucial for the parasite. All the data indicate that Albumax
I can replace human serum offering the advantages of safety, standardization
for chemosensitivity tests, and exoantigen purification.
Purification and production of Plasmodium falciparum zygotes from in vitro culture using magnetic column and Percoll density gradient
