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Background  Suitable and scalable  in vitro  culture conditions for parasite maintenance are needed to foster drug research for loiasis, one of the neglected tropical diseases which has attracted only limited attention over recent years, despite having important public health impacts. The present work aims to develop adequate  in vitro  culture systems for drug screening against both microfilariae (mf) and infective third-stage larvae (L3) of  Loa loa .    Methods   In vitro  culture conditions were evaluated by varying three basic culture media: Roswell Park Memorial Institute (RPMI-1640), Dulbecco’s modified Eagle’s medium (DMEM) and Iscove’s modified Dulbecco’s medium (IMDM); four sera/proteins: newborn calf serum (NCS), foetal bovine serum (FBS), bovine serum albumin (BSA) and the lipid-enriched BSA (AlbuMax® II, ALB); and co-culture with the Monkey Kidney Epithelial Cell line (LLC-MK2) as a feeder layer. The various culture systems were tested on both mf and L3, using survival (% motile), motility (T 90  = mean duration (days) at which at least 90% of parasites were fully active) and moulting rates of L3 as the major criteria. The general linear model regression analysis was performed to assess the contribution of each variable on the viability of  Loa loa  L3 and microfilarie. All statistical tests were performed at 95% confidence interval.    Results  Of the three different media tested, DMEM and IMDM were the most suitable sustaining the maintenance of both  L. loa  L3 and mf. IMDM alone could sustain L3 for more than 5 days (T 90  = 6.5 ± 1.1 day). Serum supplements and LLC-MK2 co-cultures significantly improved the survival of parasites in DMEM and IMDM. In co-cultures with LLC-MK2 cells,  L. loa  mf were maintained in each of the three basic media (T 90  of 16.4–19.5 days) without any serum supplement. The most effective culture systems promoting significant moulting rate of L3 into L4 (at least 25%) with substantial maintenance time were: DMEM + BSA, DMEM + NCS, DMEM-AlbuMax®II, DMEM + FBS all in co-culture with LLC-MK2, and IMDM + BSA (1.5%), DMEM + FBS (10%) and DMEM + NCS (5%) without feeder cells. DMEM + 1% BSA in co-culture scored the highest moulting rate of 57 of 81 (70.37%). The factors that promoted  L. loa  mf viability included feeder cells ( β  = 0.490), both IMDM ( β  = 0.256) and DMEM (β = 0.198) media and the protein supplements NCS ( β  = 0.052) and FBS ( β  = 0.022); while for  L. loa  L3, in addition to feeder cells ( β  = 0.259) and both IMDM ( β  = 0.401) and DMEM ( β  = 0.385) media, the protein supplements BSA ( β  = 0.029) were found important in maintaining the worm motility.    Conclusions  The findings from this work display a range of culture requirements for the maintenance of  Loa loa  stages, which are suitable for developing an effective platform for drug screening.    Electronic supplementary material  The online version of this article (10.1186/s13071-018-2852-2) contains supplementary material, which is available to authorized users.

Evaluation of in vitro culture systems for the maintenance of microfilariae and infective larvae of Loa loa