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5841 v2

Page 4 of 4

AsureQuality Laboratory Services

8. CHROMATOGRAPHIC CONDITIONS

1. Mobile Phase A : 0.1% Phosphoric acid

2. Mobile Phase B : 100% Acetonitrile

3. Mobile Phase C : 80% Acetonitrile

4. Column: Phenomenex Kinetex Phenyl-Hexyl, 00F-4495-E0 (150 x 4.6mm x 2.6μm x 100Å)

5. Column Temperature: 25±2ºC

6. Retention time: 16 to 17 minutes

7. Run time: 27 minutes

8. Detector: PDA operating at 200nm (PDA scan 200 to 350nm)

9. Injection volume: 100μL

Table 2: Gradient Program

Time (min) Flow rate (mL/min) Mobile Phase A (%) Mobile Phase B (%) Mobile Phase C (%)

0.0

0.6

90

10

0

18.0

0.6

90

10

0

18.5

0.8

0

0

100

24.0

0.8

0

0

100

24.5

0.6

90

10

0

27.0

0.6

90

10

0

9. SENSITIVITY

Limit of Quantitation (LOQ) was calculated based on the lowest working standard and the

dilution factor.

LOQ = (1 x 100) / (20 x 50) = 0.1μg/100g (1ppb)

1 = 1μg/100mL lowest standard

100 = Volume (mL)

20 = 20g sample

50 = Volume (mL) loaded on immunoaffinity column

1= Final volume (mL)

Bio-02 (February 2016)

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