Average G6PD activity (U/gHb) Trinity G6PD activity (U/gHb)

Di�

eren

ce in

G6P

D a

ctiv

ity

(U/g

Hb)

+ 1.96 SD

- 1.96 SD

Mean

Pearson correlation coe�cient [0.7585]

Linear fit equation: y = 3.84 +0.89xR2 = 0.575

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R&D

G6P

D a

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(U/g

Hb)

20

18

16

14

12

10

8

6420

4

2

0

-2

6

8

10

conclusions• AcomparisonoftwoquantitativeG6PDtestsshows:

– BothtestsequallyclassifysevereG6PDdeficient samples.

– Referencevaluesvarywidelydependingonthereferencetestused.

• TheBinaxNOW®testaccuratelydetectedallseverelydeficientpatients.

• ThefluorescentspottestonlyaccuratelydetectsallseverelydeficientspecimensifintermediatesareconsideredasG6PDdeficient.

• Bycytochemicalstaining,itwaspossibletodetectheterozygousfemalesinthepopulation.Onlythreeofthesewouldhavebeenexcludedfromtreatmentusingamoderatelydeficientcut-offof60%normalactivity.

w w w . p a t h . o r g

introductionTreatmentwithanti-malarialdrugsinthe8-aminoquinolonegroup(e.g.,primaquine,pamaquineandtafenoquine)cancauseacutehemolysisinpeoplewithglucose-6-phosphatedehydrogenase(G6PD)deficiency.Assuch,thereisaneedforatestthatisappropriateforG6PDdeficiencyscreeninginordertoensuresafeadministrationofthesedrugsforradicalcureofPlasmodium vivax.

WeevaluateddifferentassaysandplatformsfordeterminingtheG6PDstatusofapatient.

methodsAssays used to measure G6PD activity Trinity Biotech spectrophotometer test:AquantitativekineticassayexpressingG6PDactivityinunitspergramofhemoglobin.

R&D Diagnostics Ltd® enzymatic colorimetric method:AquantitativekineticcolorimetricassayexpressingG6PDactivityinunitspergramofhemoglobin.

Trinity Biotech Fluorescent spot test:Mostcommonqualitativeassay,inwhichfluorescenceofnicotinamideadeninedinucleotidephosphate(NADPH)isdirectly observed.

BinaxNOW® G6PD Test:Aqualitativeassaythatusesalateralflowtestplatform.

Cytofluorometric assay:Allowsobservationofmosaicredbloodcellpopulationsinfemales.

evaluation of diagnostic platforms for g6pd deficiency Nicole LaRue, Maria Kahn, Michael Kalnoky, Brandon T. Leader, Pooja Bansil, Sarah McGray, Gonzalo J. Domingo

table 1: trinity Biotech reference Summary of g6pD activity (N = 214)

reference values total Female Male adjusted male

Number of cases 214 107 107 101Mean (U/g Hb) 7.17 7.70 6.63 6.99Standard deviation (U/g Hb) 7.295 7.69 7.06 7.18Median (U/g Hb) 7.295 7.69 7.06 7.18

Range (U/g Hb) 0.12–14.04 1.03–14.04 0.12–12.26 0.84–12.26

table 2: Number and percent distribution of BinaxNow® and trinity Biotech Fluorescent Spot test results (N = 214)

Number (N) percent (%)

BinaxNow® test

Normal (N=201) 182 90.5

Deficient (N=201) 19 9.45

Untreated (N=214) 13 6.07

trinity Biotech Fluorescent Spot test (FSt)

Normal 189 88.32

Deficient 13 6.07

Intermediate 12 5.61

Deficient + intermediate 25 1.68

Normal + intermediate 201 93.93

figure 2: Comparison of g6pD activity classified samples by the trinity Biotech quantitative assay to two qualitative tests. the shading in the table indicates discrepancies in definition of phenotype between the tests within intermediate ranges.

Classification by trinity Biotech Quantitative

trinity Biotech Fluorescent Spot

testBinaxNow®

trinity Biotech Quantitative

Calculated activity

Sample #147 (female) NORMAL 7.50

Sample #141 (male) NORMAL 8.24

Sample #110 (female) INTERMEDIATE

3.40

Sample #105 (male) DEFICIENT 0.12

Sample #92 (male) DEFICIENT

0.66

Sample #145 (male) DEFICIENT

0.69

figure 1: pearson’s correlation coefficient and a Bland altman scatterplot describing the relationship between the trinity Biotech quantitative and r&D Diagnostics Ltd® quantitative test for g6pD activity.

results

figure 3: g6pD zygosity measured by mean FItC fluorescence compared to g6pD activity measured by trinity Biotech.

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650

10% 20% 30% 60%

Mea

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G6PD Zygosity measured by mean FITC Fluorescence and Trinity Zygosity determined by Flow Cytometry & Percent Normal determined by Trinity

G6PD activity measured by trinity (U/(g/dl)Hb)

NormalHeterozygousDeficient60% normal30% normal20% normal10% normal

results continued results continued

We thank Amanda Vilbrandt and Scott Brown for their support in the development of this poster. This poster is based on research funded by the Bill & Melinda Gates Foundation. For further information please contact nlarue@path.org, PATH, Seattle, WA.

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table 3: Clinical performance of the BinaxNow® and trinity Biotech Fluorescent Spot screening tests for detection of deficient g6pD activity*

Cut-off value (% of adjusted normal male value, 7.18)

10% 20% 30% 60%

Cut-off value (U/gHb) 0.718 1.436 2.154 4.308

Number of samples with G6PD levels below cut-off (%)

6 (2.80) 16 (7.48) 18 (8.41) 23 (10.75)

BinaxNow g6pD test

Sensitivity % (95%Cl) 100.0 (54.1, 100.0)

100.0 (79.4, 100.0)

100.0 (81.5, 100.0)

82.6 (61.2, 95.0)

Specificity % (95%Cl) 93.3 (88.9, 96.4)

98.4 (95.3, 99.7)

99.5 (97.0, 100.0)

100.0 (97.9, 100.0)

trinity Fluorescent Spot testa

Sensitivity % (95%Cl) 100.0 (54.1, 100.0)

100.0 (79.4, 100.0)

100.0 (81.5, 100.0)

91.3 (72.0, 98.9)

Specificity % (95%Cl) 90.9 (86.1, 94.4

95.5 (91.5, 97.9)

96.4 (92.8, 98.6)

97.9 (94.7, 99.4)

trinity Fluorescent Spot testb

Sensitivity % (95%Cl) 83.3 (35.9, 94.6)

75.0 (47.6, 92.7)

72.2 (46.5, 90.3)

56.5 (34.5, 76.8)

Specificity % (95%Cl) 96.2 (92.6, 98.3)

99.5 (97.2, 100.0)

100.0 (98.1, 100.0)

100.0 (98.1, 100.0)

*Gold standard: Trinity Biotech test a Trinity Biotech spot test intermediate test results combined with deficient test resultsb Trinity Biotech spot test intermediate test results combined with normal test results

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