1921
Volume 100, Issue 3
  • ISSN: 0002-9637
  • E-ISSN: 1476-1645

Abstract

Abstract.

Gametocytes are sexual stage malaria parasites responsible for transmission to mosquitoes. Multiple gametocyte-producing clones may be present in natural infections, but the molecular characterization of gametocytes is challenging. Because of their magnetic properties, gametocyte enrichment can be achieved by magnetic fractionation. This increases detection sensitivity and allows specific genotyping of clones that contribute to malaria transmission. Here, we determined the percentage of gametocytes successfully bound to magnetic activated cell sorting (MACS) LS columns during magnetic fractionation and assessed whether columns can be reused without risking contamination or affecting column binding efficiency. Bound column fractions were quantified using multiplex quantitative reverse transcription polymerase chain reaction (qRT-PCR) for male (pfMGET) and female (CCp4) gametocytes and ring-stage asexual parasites (SBP1). To investigate cross contamination between columns, parasite strain identity was determined by merozoite surface protein 2 genotyping followed by capillary electrophoresis fragment sizing. A reproducible high percentage of gametocytes was bound to MACS LS columns with < 5% gametocytes appearing in the flow-through and < 0.6% asexual ring-stage parasites appearing in the gametocyte fraction. A high yield (> 94%) of gametocyte enrichment was achieved when columns were used up to five times with lower binding success after eight times (79%). We observed no evidence for cross contamination between columns.

[open-access] This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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  • Received : 24 Sep 2018
  • Accepted : 26 Oct 2018
  • Published online : 02 Jan 2019

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