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Table 3 Impact of stochastic variation when estimating parasite density from thick or thin films with nominated parasite density. Results from simulations of counting parasites on thick films against 200 WBC and 500 WBC (assuming 8000 WBC per µL; (A)) or on a thin film per 1000 RBC and 2000 RBC (assuming 5 × 106 RBC per µL; (B)). Parasite density estimates obtained from a single read by counting parasites per 500 WBC or 2000 RBC tend to be closer to the actual density. When the parasite density is estimated from two readings, fewer paired reads are discordant per Obare criteria [30]. Parasite density estimates were defined as being discordant if there is a < 10% chance of observing the two read densities if both were random samples from the theoretical probability distribution with mean equal to the average of the first and second read densities, given that at least one of the first or second reads has a density for a particular species above 200 parasites/µL

From: Towards harmonization of microscopy methods for malaria clinical research studies

  True parasite density (parasites/µL)
200 250 500 1000 5000 10,000 16,000
(A) Thick film counts
Against 200 WBC
Distribution of single read (n = 1000)
Mean (sd) 199.8 (89.1) 250.8 (101.6) 496.7 (138.9) 983.5 (200.1) 5018.5 (565.1) 10,041.7 (904.6) 16,001.9 (1357.9)
25th and 75th percentiles 120 and 261 189 and 314 396 and 585 839 and 1121 4619 and 5397 9412 and 10,624 15,080 and 16,956
5th and 95th percentiles 76 and 355 82 and 429 272 and 731 673 and 1320 4121 and 5989 8610 and 11,604 13,822 and 18,339
Simulation of 2 microscopy readings (n = 10,000)  
% with both readings ≤ 200/uL 34.3 13.9 0.02 0.0 0.0 0.0 0.0
% requiring 3rd read (Obare criteria) 12.7 15.4 15.3 15.4 23.1 28.7 36.9
Against 500 WBC
Distribution of single read (n = 1000)
Mean (sd) 199.7 (57.0) 248.9 (64.5) 499.5 (89.2) 998.3 (133.8) 5004.5 (354.2) 9995.1 (579.5) 15,964.0 (883.0)
25th and 75th percentiles 158 and 238 204 and 287 441 and 559 910 and 1085 4756 and 5247 9607 and 10,372 15,395 and 16,556
5th and 95th percentiles 112 and 300 144 and 365 351 and 648 780 and 1213 4441 and 5605 9076 and 10,992 14,534 and 17,524
Simulation of 2 microscopy readings (n = 10,000)  
% with both readings ≤ 200/µL 25.3 4.9 0.0 0.0 0.0 0.0 0.0
% requiring 3rd read (Obare criteria) 1.9 3.0 2.5 2.8 5.5 9.8 16.9
  True parasite density (parasites/µL)
16,000 20,000 32,000 64,000 128,000 200,000 250,000
(B) Thin film counts
Against 1000 RBC
Distribution of single read (n = 1000)
Mean (sd) 16,328.4 (8892.7) 19,979.2 (9541.3) 32,539.0 (12,592.1) 63,489.3 (17,600.1) 127,011.1 (23,853.1) 200,712.7 (32,038.7) 252,236.6 (33,445.7)
25th and 75th  %iles 9609 and 22,386 13,915 and 24,919 23,747 and 39,969 51,450 and 75,794 110,391 and 142,273 179,076 and 221,459 229,845 and 274,556
5th and 95th  %iles 4617 and 33,049 4818 and 37,912 14,020 and 53,447 34,967 and 93,502 89,799 and 168,045 147,499 and 254,466 194,879 and 306,865
Simulation of 2 microscopy readings (n = 10,000)
% requiring 3rd read (Obare criteria) 11.5 12.1 13.6 14.5 11.9 14.5 10.6
Against 2000 RBC
Distribution of single read (n = 1000)
Mean (sd) 16,116.9 (6051.3) 20,140.1 (6869.9) 3773.0 (9055.7) 64,819.4 (12,493.7) 128,216.7 (17,857.2) 199,747.5 (22,125.0) 249,103.6 (25,238.4)
25th and 75th  %iles 11,872 and 19,666 15,956 and 24,070 25,668 and 37,926 56,610 and 72,317 115,908 and 140,332 183,871 and 215,118 231,741 and 265,957
5th and 95th  %iles 7076 and 26,527 9397 and 31,699 18,248 and 47,561 45,282 and 86,739 99,791 and 159,231 164,356 and 235,178 209,136 and 291,824
Simulation of 2 microscopy readings (n = 10,000)
% requiring 3rd read (Obare criteria) 2.5 3.1 4.6 3.7 3.5 3.3 3.5