Ractopamine Elisa Kit for Food safety,96T

Ractopamine Elisa Kit-Food Safety

INTENTED USE

Ractopamine ELISA Kit is an indirect enzyme-linked immunosorbent assay for the quantitative detect the presence of ractopamine in samples of animal’s tissue, urine and feed.

Sensitivity: 0.025 ppb

Detection Limit: Animal’s tissue 0.1 ppb / 0.025 ppb

Urine 0.025 ppb

Feed 1.25 ppb

Recovery Rate: 90% ± 15% (pork liver, feed 80% ± 15%)

KIT COMPONENT

• 1 X Microtiter plate (8 microwells X 12 removable strips)
• 6 X Standard solutions (1 mL each): 0 ng/ mL, 0.025 ng/ mL, 0.075 ng/ mL, 0.225 ng/ mL, 0.675 ng/ mL, 2.025 ng/ mL
• 1 X Ractopamine antibody (6 mL)
• 1 X HRP conjugated antibody (6 mL)
• 1 X Sample dilution buffer (30 mL)
• 1 X Washing buffer concentrate (10 X , 50 mL)
• 1 X Substrate A (6 mL)
• 1 X Substrate B (6 mL)
• 1 X Stopping solution (6 mL)
• 1 X Product Manual

1. MATERIALS REQUIRED BUT NOT PROVIDED

• ELISA Microtiter plate reader equipped with 450 nm filters

- 50-200 μL multichannel micropipette

- 50, 100 and 200 μL precision micropipette

- Microplate washer or squeeze bottle

- Homogenizer

- Centrifuge

- Centrifugal tubes

- Distilled water

- HCl, NaOH

PREPARATION OF WORKING SOLUTIONS

Washing buffer: dilute 10X concentrate with distilled water (i.e. 1 mL + 9 mL).

1 M HCl: dilute 8.33mL concentrated hydrochloric acid with distilled water and make the constant volume to 100 mL, OR dilute 10X 1M HCl with distilled water (i.e. 1 mL + 9 mL).

Extract solution: mix 8g NaCl with 100mL 1M HCl .

Standard solution, Enzyme-linked conjugate, substrate A, substrate B and stopping solution are ready-to-use.

SAMPLE PREPARATION

1.Animal’s tissue

Method 1 (dilution factor: 4):

• Homogenize the sample (free of fat) for 1 min at 10,000 r/min.
• Weigh out 2.0 g of homogenate into a 15 mL centrifugal tube. Add 6 mL of extract solution and mix well and shake for 5 min.
• Do a centrifugation at 5000 rpm at 20-25 °C for 5 min.
• Take 1 mL of supernatant to polystyrene centrifuge tube and mix it with 20 μL of 1 M NaOH. Adjust the pH value to 7.0-8.0.
• Take 50 μL liquid from water phase for assay.

Method 2 (dilution factor: 1):

• Homogenize the sample (free of fat) for 1 min at 10,000 r/min.
• Weigh out 2.0 g of homogenate into a 15 mL centrifugal tube. Add 6 mL of Methyl alcohol and shake for 5 min.
• Do a centrifugation at 5000 rpm at 20-25 °C for 5 min.
• Take 1.5 mL of supernatant (organic layer) into a glass tube and dry it by 50 °C of Nitrogen flow.
• Dissolve the residue by 0.5 mL of sample dilution buffer and add 0.5 mL of n-Hexane. Shake to mix well.
• Transfer 1ml into centrifuge tube.
• Get rid of the upper layer and take 50 μL liquid for assay.
• 2.Urine (dilution factor 1)
• Take 50 μL of clear urine sample directly for assay. Make dilution by distilled water before assay if the concentration of ractopamine is high.
• 3.Feed (dilution factor 50)
• Weigh out 2 g of comminuted feed sample into a 50 mL centrifugal tube. Add 2 mL of 1 M HCl and 16 mL of distilled water. Add 1.6g NACL, mix well.
• Do a centrifugation at 5000 rpm at 20-25 °C for 5 min.
• Take 200 μL of supernatant and mix well with 800 μL of sample dilution buffer. Shake to mix well.
• Adjust the pH value to 7.0-8.0 by 1 M NaOH.

Take 50 μL for assay.