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The lower limit of microscopy for gametocyte quantification was therefore estimated at 8 gametocytes/ µl of blood.
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There was a strong correlation between gametocyte densities detected by QT-NASBA and microscopy for microscopically gametocyte positive samples (Spearman correlation coefficient = 0.60; p = 0.004).
Venous blood samples (3 mL) were drawn into heparin-containing tubes for membrane feeding and for gametocyte detection both by microscopy and by real-time Pfs25 quantitative nucleic acid sequence based amplification (QT-NASBA).
When the Pfs25 QT-NASBA was used for gametocyte detection, 91.6% (65/71) individuals were shown to be carrying gametocytes.
For gametocyte traits with repeated measures for the same individual (i.e. gametocyte positivity and gametocyte density), a Generalized Linear Mixed Model (GLMM) was fitted with individual person as a factor in the random model.
Person-gametocyte-weeks (PGW expressed per 1000) were defined as the number of weeks in which blood slides were positive for gametocyte divided by the total number of weeks followed up in patients with gametocyte results [19].
This would result in a smaller source of asexual parasites for gametocyte production and hence a reduced gametocyte density that makes their detection more difficult.
For gametocyte positivity, the impact of these factors was, however, small.
Asexual parasite density was estimated by microscopy while gametocyte density was estimated using QT-NASBA.
To determine, if EM permeabilization is required for gametocyte egress we treated gametocytes with the pore sealant Tetronic 90R4 prior to their activation.
Subsequently, we evaluated if EM permeabilization is a prerequisite for gametocyte egress from the erythrocyte.
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