Analytical Performance Evaluation Report for

HbA1c Rapid Quantitative Test

Guangzhou Wondfo Biotech Co., Ltd had started the research and development, trial production
and laboratory evaluation of Finecare HbA1c Rapid Quantitative Test (Immunofluorescence
Method) from 2011. This documents was drawn up base on the evaluation results above, which
followed the Product Quality Control Regulation Standard of Chinese Biological Products
Procedure (Version 2000) drafted by Committee for Biological Product Standardization in China.

Content 
The lowest detection limit study ....................................................................................... 2 
Linear Study ...................................................................................................................... 3 
Reportable range ............................................................................................................... 6 
Accuracy study (Methodology comparison) ..................................................................... 9 
Precision study ................................................................................................................ 11 
Conclusion ...................................................................................................................... 14 
The lowest detection limit study

1. Bank sample: 0.5%BSA solution

2. Test method: Take 3 batches of HbA1c rapid quantitative test cartridges, test each batch with

blank sample for 20 times. Record the results as below, use ( X +2SD) to record the lowest

detection limit.
3. Test result:
Table 1 The lowest detection limit
Lot No.
W207B301A W207B302A W207B303A
Test No.

1 3.5% 3.5% 3.6%

2 3.7% 3.5% 3.6%

3 3.5% 3.6% 3.7%

4 3.2% 3.4% 3.4%

5 3.2% 3.4% 3.3%

6 3.4% 3.2% 3.5%

7 3.6% 3.8% 3.6%

8 3.0% 3.5% 3.7%

9 3.4% 3.0% 3.3%

10 2.9% 3.6% 3.1%

11 3.0% 3.4% 3.0%

12 2.9% 3.1% 3.8%

13 3.4% 3.1% 3.6%

14 3.4% 3.6% 3.5%

15 3.2% 3.7% 3.4%

16 3.6% 3.4% 3.4%

17 3.6% 3.2% 3.3%

18 3.5% 3.3% 3.2%

19 3.0% 3.3% 3.1%

20 3.3% 3.5% 3.1%

 SD 0.0025 0.0021 0.0023

X 0.0332 0.0341 0.0341

X +2SD 3.8% 3.8% 3.9%

4. Conclusion: ( X +2SD) is ＜4%, we take 4% as the lowest detection limit.

Linear Study

1. Sample: Low concentration HbA1c blood sample (5.3%), high concentration HbA1c blood
sample (13.9%). Add some substrate to make the low concentration HbA1c blood sample
concentration reduces to 4%, and add some pure HbA1c to high concentration HbA1c blood
sample concentration rise to 14.5%. Mix the samples by different volume proportion as
below:
Table 2 Linear samples preparation
Sample NO. 1 2 3 4 5 6 7 8 9 10 11

Volume of low concentration

1.00 0.90 0.80 0.70 0.60 0.50 0.40 0.30 0.20 0.10 0.00
sample（ml）

Volume of high concentration

0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80 0.90 1.00
sample（ml）

2. Method
Take 3 batches of HbA1c rapid quantitative test cartridges to test the sample of above samples of
11 levels, each sample should repeat test for 4 times. Record the results and calculate the
correlation as below.
Table 3 Linear data record
Result Lot NO. W207B301A

Sample No. 1 2 3 4 Mean

1 4.0% 4.0% 4.0% 4.0% 4.0%

2 5.4% 4.9% 5.3% 5.0% 5.2%

3 6.0% 6.3% 6.0% 6.4% 6.2%

4 7.4% 7.3% 7.4% 7.4% 7.4%

5 8.0% 8.3% 8.2% 8.1% 8.2%

6 9.5% 9.4% 9.5% 9.5% 9.5%

 7 10.6% 10.5% 10.6% 10.4% 10.5%

8 11.1% 11.0% 10.9% 10.9% 11.0%

9 12.0% 11.8% 11.6% 11.9% 11.8%

10 13.5% 13.6% 13.4% 13.0% 13.4%

11 14.8% 14.9% 14.5% 15.0% 14.8%

r 0.9970

Regression
y = 0.0103x + 0.0307
equation

16.0%
y = 0.0103x + 0.0307
14.0%
R² = 0.9941
12.0%

10.0%
系列1
8.0%
线性 (系列1)
6.0%

4.0%

2.0%

0.0%
0 5 10 15

Table 4 Linear data record

Result Lot No. W207B302A

Sample NO. 1 2 3 4 Mean

1 4.0% 4.0% 4.0% 4.0% 4.0%

2 4.9% 4.8% 4.8% 4.8% 4.8%

3 6.3% 6.2% 6.3% 6.4% 6.3%

4 7.1% 7.0% 7.1% 6.9% 7.0%

5 8.3% 8.4% 8.3% 8.2% 8.3%

6 9.0% 9.1% 8.9% 8.8% 9.0%

7 10.3% 10.5% 10.6% 10.7% 10.5%

8 11.5% 11.6% 11.5% 11.6% 11.6%

9 12.2% 12.3% 12.3% 12.1% 12.2%

 10 14.0% 14.0% 13.8% 13.7% 13.9%

11 14.5% 14.6% 14.5% 14.2% 14.5%

r 0.9981

Regression
y = 0.0107x + 0.0287
equation

16.0%

y = 0.0107x + 0.0287
14.0% R² = 0.9963

系列1
12.0%
线性 (系列1)

10.0%

8.0%

6.0%

4.0%

2.0%

0.0%
0 2 4 6 8 10 12

Table 5 Linear data record

Result Lot No. W207B303A

Sample No. 1 2 3 4 Mean

1 4.0% 4.0% 4.0% 4.0% 4.0%

2 5.2% 5.3% 5.4% 5.1% 5.3%

3 6.0% 6.0% 6.2% 6.0% 6.1%

4 7.6% 7.4% 7.4% 7.5% 7.5%

5 8.3% 8.5% 8.6% 8.4% 8.5%

6 9.2% 9.3% 9.2% 9.1% 9.2%

7 10.0% 9.9% 10.1% 9.9% 10.0%

8 11.5% 11.4% 11.6% 11.6% 11.5%

9 12.1% 11.9% 12.0% 11.8% 12.0%

10 14.0% 13.9% 13.8% 13.7% 13.9%

11 14.4% 14.6% 14.5% 14.2% 14.4%

 r 0.9971

Regression y = 0.0104x + 0.0308

equation

16.0%
y = 0.0104x + 0.0308
14.0% R² = 0.9943
系列1
12.0%
线性 (系列1)

10.0%

8.0%

6.0%

4.0%

2.0%

0.0%
0 2 4 6 8 10 12

3. Conclusion
Above results indicate that HbA1c Rapid Quantitative Test has good linear correlation in
concentration of 4.0%～14.5%, the correlation coefficient ≥0.99.

Reportable range

1. Sample selection
Low value sample: take 5 reference samples with HbA1c theoretical value of 2%, 3%, 4%, 5%
and 6%
High value sample: take 5 reference samples with HbA1c theoretical value of 14%, 14.5%
and 15%

2. Method
Take 3 batches of HbA1c rapid quantitative test cartridges（lot No.：W207B301A，
W207B302A and W207B303A） to test the sample of above samples. Each low value sample
should repeat test for 10 times, each high value sample should be diluted by matrix and repeat
test for 3 times. Record the results and calculate the correlation as below

3. Result
The acceptable low limit for reportable range base on CV≤10.0%, choose the lowest value in
data sheet that conform to CV≤10.0%. The acceptable high limit for reportable range base on
relative deviation≤±10.0%, choose the highest value in data sheet that conform to relative
deviation≤±10.0%.
Table 6 Reportable range data record (limit down)
 Theoretical Lot No.：W207B301A

value
2% 3% 4% 5% 6%
Test NO.

1 2.2% 2.7% 4.0% 5.6% 5.7%

2 1.8% 2.7% 4.2% 5.6% 5.5%

3 2.0% 2.8% 3.7% 5.4% 5.8%

4 2.0% 2.8% 4.6% 5.2% 6.2%

5 2.3% 2.6% 4.1% 5.3% 6.0%

6 2.4% 2.6% 3.8% 4.9% 6.1%

7 2.5% 3.0% 4.0% 5.4% 6.3%

8 2.4% 3.2% 3.6% 5.5% 6.4%

9 1.9% 3.3% 3.9% 4.8% 6.3%

10 1.8% 3.5% 4.5% 5.2% 6.2%

Mean 2.1% 2.9% 4.0% 5.3% 6.1%

SD 0.0026 0.0032 0.0032 0.0027 0.0030

CV% 12.33% 10.81% 8.02% 5.15% 4.88%

Theoretical Lot No.：W207B302A

value
2% 3% 4% 5% 6%
Test NO.

1 2.3% 2.6% 4.0% 4.9% 6.2%

2 2.2% 2.6% 3.6% 5.3% 6.3%

3 2.0% 2.6% 3.7% 5.5% 6.0%

4 2.2% 2.7% 3.6% 5.6% 6.7%

5 2.4% 2.8% 3.8% 4.7% 5.8%

6 2.3% 3.0% 4.0% 5.2% 5.9%

7 1.7% 3.4% 4.5% 4.8% 5.8%

8 1.8% 2.8% 4.4% 5.0% 6.7%

9 1.8% 3.2% 4.3% 4.7% 6.3%

10 1.9% 3.4% 4.4% 5.1% 6.5%

 Mean 2.06% 2.91% 4.03% 5.08% 6.22%

SD 0.0025 0.0032 0.0035 0.0032 0.0034

CV% 12.15% 11.04% 8.68% 6.28% 5.51%

Theoretical Lot No.：W207B303A

value
2% 3% 4% 5% 6%
Test NO.

1 2.1% 3.3% 4.0% 5.0% 6.6%

2 1.8% 3.4% 3.8% 5.5% 6.5%

3 1.7% 3.0% 3.6% 5.4% 6.4%

4 1.6% 3.0% 4.4% 5.1% 6.2%

5 2.0% 3.4% 4.3% 4.8% 6.0%

6 2.2% 2.7% 4.2% 4.6% 5.8%

7 2.1% 2.6% 4.4% 4.9% 5.9%

8 2.2% 2.8% 4.0% 5.0% 5.7%

9 1.8% 2.6% 3.7% 4.7% 6.2%

10 1.7% 3.3% 3.8% 5.3% 6.0%

Mean 1.92% 3.01% 4.02% 5.03% 6.13%

SD 0.0023 0.0032 0.0029 0.0030 0.0030

CV% 11.72% 10.79% 7.30% 5.93% 4.93%

Table 7 Reportable range data record (limit up)

Theoretical value Lot No.：W207B301A

Test NO. 14% 14.5% 15%

1 4.4% 4.3% 4.5%

2 4.2% 4.4% 4.4%

3 4.1% 4.4% 4.4%

Mean 4.2% 4.4% 4.4%

Dilution ratio 3 3 3

Restore Conc. 12.7% 13.1% 13.3%

 Relative
9.29% 9.66% 11.33%
deviation（%）

Theoretical value Lot No.：W207B302A

Test NO. 14% 14.5% 15%

1 4.2% 4.4% 4.4%

2 4.3% 4.4% 4.4%

3 4.3% 4.4% 4.6%

Mean 4.3% 4.4% 4.5%

Dilution ratio 3 3 3

Restore Conc. 12.8% 13.2% 13.4%

Relative
8.57% 8.97% 10.67%
deviation（%）

Theoretical value Lot No.：W207B303A

Test NO. 14% 14.5% 15%

1 4.3% 4.3% 4.3%

2 4.3% 4.3% 4.5%

3 4.3% 4.5% 4.3%

Mean 4.3% 4.4% 4.4%

Dilution ratio 3 3 3

Restore Conc. 12.9% 13.1% 13.1%

Relative
7.86% 9.66% 12.67%
deviation（%）

4. Conclusion
Above results indicate that for the low value samples (theoretical concentration of 2%~6%), 4%
is lowest value with CV≤10.0%, we set it as limit down. For the high value samples
(theoretical concentration of 14%~15%), 14.5% is lowest value with relative deviation≤±
10%, we set it as limit up. The reportable range is 4.0%～14.5%.

Accuracy study (Methodology comparison)

1. Test reagent/test device: Finecare HbA1c Rapid Quantitative Test (Immunofluorescence
 Method) and Finecare FIA Meter manufactured by Guangzhou Wondfo Biotech Co., Ltd., lot
number: W207B301A.
2. Contrast reagent/test device: HbA1c Test Kit (HPLC Method) and D-10 HbA1c Analyzer
manufactured by American BioRad Company.
3. Sample source: 8 HbA1c samples per day provided by Jiangmen Wuyi Hospital of TCM,
continuously tested for 5 days by both Wondfo test and BioRad tests.
4. Test method: Follow the instructions, compare the test correlations.
5. Result: record the results, draw the mean scatter diagram.
Table 8 Accuracy test result
Day 1 Day 2 Day 3 Day 4 Day 5

NO. BioRad Wondfo NO. BioRad Wondfo NO. BioRad Wondfo NO. BioRad Wondfo NO. BioRad Wondfo

1 8.3% 8.5% 9 5.8% 5.6% 17 4.3% 4.2% 25 11.2% 11.3% 33 4.7% 4.7%

2 5.2% 5.0% 10 10.6% 10.5% 18 4.0% 4.3% 26 10.2% 10.6% 34 4.6% 4.6%

3 5.0% 5.4% 11 5.9% 6.0% 19 6.0% 6.3% 27 7.1% 7.1% 35 4.2% 4.6%

4 4.9% 4.9% 12 5.0% 5.3% 20 7.3% 7.0% 28 6.8% 6.6% 36 9.2% 9.4%

5 4.8% 4.4% 13 5.4% 5.7% 21 7.1% 7.1% 29 12.0% 11.6% 37 5.5% 5.1%

6 4.6% 4.6% 14 9.3% 9.2% 22 6.6% 6.2% 30 10.2% 10.6% 38 5.2% 5.3%

7 4.5% 4.3% 15 5.8% 6.0% 23 5.5% 5.1% 31 4.0% 4.2% 39 5.4% 5.0%

8 5.3% 5.1% 16 8.6% 9.0% 24 4.0% 4.3% 32 4.0% 4.3% 40 4.8% 5.0%
 14.0%
y = 0.9971x + 0.0005
12.0%
R² = 0.9855
10.0%

8.0%
系列1
6.0% 线性 (系列1)

4.0%

2.0%

0.0%
0.0% 5.0% 10.0% 15.0%

6. Conclusion:
The test result of Wondfo Finecare HbA1c Rapid Quantitative Test (Immunofluorescence Method)
is accurate and reliable. The correlation between Wondfo HbA1c Rapid Quantitative Test and
BioRad HbA1c Test Kit (HPLC Method) is good, correlation coefficient r = 0.9927, regression
equation = 0.9971x+0.0005.

Precision study
1. Sample: HbA1c reference samples concentration of 7.8% and 12.3%.
2. Method: take 3 batches of Finecare HbA1c Rapid Quantitative Test (Lot: W207B301A,
W207B302A and W207B303A) to test HbA1c reference samples concentration of 7.8% and 12.3%
respectively. Each sample should be tested for 8 times per day for each batch, 5 days experiment
in total.
3. Result: Calculate within-run precision with 40 test results from one batch, calculate
between-run precision with 120 test results from the 3 batches.
Table 9 Precision test result record (lot W207B301A, HbA1c conc. 7.8%)
Result Lot：W207B301A (HbA1c concentration：7.8%)

Days 1 2 3 4 5 6 7 8 Mean SD

1 7.6% 7.4% 7.3% 7.3% 7.4% 7.1% 7.9% 8.0% 7.5% 0.0031

2 7.0% 7.1% 7.3% 7.0% 7.0% 7.2% 7.5% 7.4% 7.2% 0.0020

3 7.2% 7.3% 7.2% 7.0% 8.1% 7.9% 7.8% 7.9% 7.6% 0.0042

4 7.1% 7.3% 7.3% 7.5% 7.4% 7.3% 7.1% 7.3% 7.3% 0.0014

5 7.3% 7.5% 7.4% 7.1% 7.9% 7.0% 7.3% 7.8% 7.4% 0.0031

Mean 7.4%
 SD 0.3%

CV% 3.7%

Table 10 Precision test result record (lot W207B302A, HbA1c conc. 7.8%)
Result Lot：W207B302A (HbA1c concentration：7.8%)

Days 1 2 3 4 5 6 7 8 Mean SD

1 7.0% 7.0% 6.9% 7.1% 7.2% 7.5% 7.4% 7.9% 7.3% 0.0033

2 7.3% 7.1% 8.2% 8.4% 7.8% 7.9% 7.3% 8.0% 7.8% 0.0047

3 7.4% 7.5% 7.5% 7.8% 7.8% 7.9% 8.4% 8.1% 7.8% 0.0034

4 7.6% 7.0% 7.1% 7.5% 7.4% 7.4% 8.0% 8.0% 7.5% 0.0037

5 7.6% 7.7% 7.4% 7.5% 7.8% 7.7% 8.2% 7.7% 7.7% 0.0024

Mean 7.6%

SD 0.3%

CV% 4.6%

Table 11 Precision test result record (lot W207B303A，HbA1c conc. 7.8%)

Result Lot：W207B303A (HbA1c concentration：7.8%)

Days 1 2 3 4 5 6 7 8 Mean SD

1 8.3% 8.0% 8.1% 7.6% 7.7% 7.5% 7.1% 7.4% 7.7% 0.0040

2 7.6% 8.0% 8.1% 8.2% 7.9% 7.3% 7.3% 7.4% 7.7% 0.0037

3 7.6% 7.4% 7.4% 7.3% 7.1% 7.5% 7.9% 8.3% 7.6% 0.0038

4 7.8% 7.6% 8.1% 8.3% 7.8% 7.2% 7.9% 8.0% 7.8% 0.0033

5 7.7% 7.2% 7.1% 7.8% 7.3% 7.5% 7.9% 8.0% 7.6% 0.0034

Mean 7.7%

SD 0.4%

CV% 4.7%

Table 12 Precision test result record (lot W207B301A，HbA1c conc. 12.3%)

Result Lot：W207B301A (HbA1c concentration：12.3%)

Days 1 2 3 4 5 6 7 8 Mean SD
 1 11.5% 11.6% 11.8% 12.0% 12.6% 12.3% 13.2% 12.8% 12.2% 0.0061

2 11.8% 11.8% 11.7% 12.1% 12.2% 13.0% 12.3% 12.9% 12.2% 0.0049

3 11.9% 12.3% 12.3% 12.4% 12.9% 12.8% 12.6% 13.0% 12.5% 0.0037

4 11.7% 11.8% 11.6% 12.0% 12.1% 12.1% 12.5% 12.6% 12.1% 0.0036

5 12.5% 12.4% 12.3% 12.8% 12.1% 11.8% 11.9% 12.6% 12.3% 0.0035

Mean 12.3%

SD 0.4%

CV% 3.5%

Table 13 Precision test result record (lot W207B302A，HbA1c conc. 12.3%)

Result Lot：W207B302A (HbA1c concentration：12.3%)

Days 1 2 3 4 5 6 7 8 Mean SD

1 12.0% 12.3% 12.6% 12.8% 13.0% 11.8% 11.9% 12.5% 12.4% 0.0044

2 12.4% 12.9% 12.7% 12.8% 13.1% 13.0% 11.9% 12.5% 12.7% 0.0039

3 12.4% 12.5% 12.8% 12.7% 12.6% 12.2% 11.8% 11.7% 12.3% 0.0041

4 11.4% 11.8% 11.7% 12.9% 12.8% 12.5% 12.4% 12.2% 12.2% 0.0054

5 12.6% 13.0% 12.9% 12.4% 12.8% 11.5% 11.6% 12.1% 12.4% 0.0058

Mean 12.4%

SD 0.5%

CV% 3.8%

Table 14 Precision test result record (lot W207B303A，HbA1c conc. 12.3%)

Result Lot：W207B303A (HbA1c concentration：12.3%)

Days 1 2 3 4 5 6 7 8 Mean SD

1 12.5% 12.4% 12.0% 11.8% 11.5% 11.4% 11.8% 12.8% 12.0% 0.0050

2 12.4% 11.6% 11.9% 11.8% 12.0% 12.4% 12.5% 12.9% 12.2% 0.0043

3 12.9% 12.3% 12.3% 12.2% 12.0% 11.8% 11.7% 12.0% 12.2% 0.0037

4 12.1% 11.8% 12.1% 12.5% 12.5% 12.6% 12.6% 13.0% 12.4% 0.0038
 5 11.9% 12.5% 12.6% 12.8% 13.2% 13.0% 12.4% 12.9% 12.7% 0.0041

Mean 12.3%

SD 0.4%

CV% 3.4%

4. From table 9 to table 14 we calculated the CV and 95% confidence limit of different test
batches and different concentration HbA1c samples, results below:
Table 15 Within-run and between-run precision result 1
HbA1c concentration：7.8%

Lot Mean SD CV% 95% Cl

W207B301A 7.4% 0.3% 3.7% 6.8% 8.0%

Within
W207B302A 7.6% 0.3% 4.6% 7.0% 8.2%
run
W207B303A 7.7% 0.4% 4.7% 6.9% 8.5%

Between run 7.6% 0.3% 4.3% 6.9% 8.2%

Table 16 Within-run and between-run precision result 2

HbA1c concentration：12.3%

Lot Mean SD CV% 95% Cl

W207B301A 12.3% 0.4% 3.5% 11.5% 13.1%

Within
W207B302A 12.4% 0.5% 3.8% 11.4% 13.4%
run
W207B303A 12.3% 0.4% 3.4% 11.5% 13.1%

Between run 12.3% 0.4% 3.6% 11.5% 13.2%

6. Conclusion: Finecare HbA1c Rapid Quantitative Test has good within-run precision and
between-run precision, the CV for both concentration HbA1c reference samples are ＜10.0%.

Conclusion

For Finecare HbA1c Rapid Quantitative Test,

Lowest detection limit is 4.0%.

Linear range is 4.0%～14.5%.

Reportable range is 4.0%～14.5%.

Accuracy: The test result of Wondfo Finecare HbA1c Rapid Quantitative Test
(Immunofluorescence Method) is accurate and reliable. The correlation between Wondfo HbA1c
Rapid Quantitative Test and BioRad HbA1c Test Kit (HPLC Method) is good, correlation
coefficient r = 0.9927, regression equation = 0.9971x+0.0005.

Precision: Finecare HbA1c Rapid Quantitative Test has good within-run precision and
between-run precision, the CV for both concentration HbA1c reference samples are ＜10.0%.