Preparation of RIGS kit.
A novel format (Fig. 1) was designed for RIGS kit, so that it enables the detection of TSWV on a single strip.
The colloidal-gold-conjugated solution of antisera raised against TSWV was mixed in equal quantities
TSWV IgG-BSA conjugate was diluted in 20 mM sodium phosphate buffer, (pH 8.0) containing 1% sucrose to a final concentration of 2 mg/ml and applied as a 0.5 mm thick, 26 cm line, centrally at 1.25 cm from the top and bottom ends on one side of a 2.5 × 26 cm Nitrocellulose membrane was cut into sections (2.5 cm × 26 cm). Test line was coated with TSWV-IgG conjugate, which was applied to each membrane in 150 μg/ml TSWV using TLC conjugate sampler.
The distance between the test line and control line was 6mm. The test strips were dried at 37°C for 30 min.
TSWV IgGBSA conjugate was diluted in 1 mM Tris-HCl buffer (pH 8.0) containing
containing 1% sucrose to a final concentration of 3 μg/ml and applied as a 1 mm thick, 26 cm line, at 0.5 cm from the top end of the membrane.
The membrane was dried at 3°C under a dry wind blower for 10-15 min and blocked with 10 mM potassium phosphate buffer (pH 7.5) containing 0.25% BSA and 1.0% sucrose.
The membrane was dried at 37°C under a dry wind blower for 10-15 min and washed twice with 10 mM,sodium phosphate buffer (pH 7.2) before drying it for 10–15 min at 50°C.
A filter pad was placed to overlap 2mm on the lower end of the conjugate release pad to act as sample pad and another pad (CF4) was placed to overlap 2 mm on the upper end of the membrane to act as absorbent pad.
The strips were stored in an airtight plastic bottle containing a desiccant pack.
Preparation of RIGS kit. A novel format (Fig. 1) was designed for RIGS kit, so that it enables the detection of TSWV on a single strip. The colloidal-gold-conjugated solution of antisera raised against TSWV was mixed in equal quantities TSWV IgG-BSA conjugate was diluted in 20 mM sodium phosphate buffer, (pH 8.0) containing 1% sucrose to a final concentration of 2 mg/ml and applied as a 0.5 mm thick, 26 cm line, centrally at 1.25 cm from the top and bottom ends on one side of a 2.5 × 26 cm Nitrocellulose membrane was cut into sections (2.5 cm × 26 cm). Test line was coated with TSWV-IgG conjugate, which was applied to each membrane in 150 μg/ml TSWV using TLC conjugate sampler. The distance between the test line and control line was 6mm. The test strips were dried at 37°C for 30 min. TSWV IgGBSA conjugate was diluted in 1 mM Tris-HCl buffer (pH 8.0) containingcontaining 1% sucrose to a final concentration of 3 μg/ml and applied as a 1 mm thick, 26 cm line, at 0.5 cm from the top end of the membrane. The membrane was dried at 3°C under a dry wind blower for 10-15 min and blocked with 10 mM potassium phosphate buffer (pH 7.5) containing 0.25% BSA and 1.0% sucrose. The membrane was dried at 37°C under a dry wind blower for 10-15 min and washed twice with 10 mM,sodium phosphate buffer (pH 7.2) before drying it for 10–15 min at 50°C. แผ่นกรองถูกวางทับ 2 มม.ในส่วนล่างของแผ่นรุ่น conjugate ทำหน้าที่เป็นแผ่นตัวอย่าง และอีกแผ่น (CF4) ถูกวางทับ 2 มม.ในส่วนบนของเมมเบรนทำหน้าที่เป็นแผ่นดูดซับ แถบถูกเก็บไว้ในขวดพลาสติกแบบสุญญากาศประกอบด้วยแพคความชื้น
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