Fumonisins伏馬毒素-定量-快篩試紙-酒精萃取

• 原理
  側流體免疫層析法 (lateral flow immunochromatographic assay)，為快篩試劑的使用原理，藉由抗體抗原間的專一性與免疫親和力做檢測，被普遍應用在、病毒感染確認、妊娠測試、警局臨檢驗毒等方面。此方法多用於定性檢測，且顯色標定物上也有許多不同的種類，目前大多數的顯色方式是利用抗體吸附在膠體金（colloid gold）來呈色。
  
  伏馬鐮孢毒素（Fumonisins)又稱伏馬毒素，此毒素主要由真菌-鐮孢菌（Fusarium）產生，伏馬毒素B₁+B₂是最常被檢出的種類，其檢出含量也常最高，約佔總伏馬毒素量之80到90%，故B₁ 和 B₂ 為伏馬毒素群中最具代表性之毒素。此毒素常出現於飼料或食物原料(例如:小麥、米類、玉米)，且會造成馬腦白質軟化症（Equine leukoence phalomalacia, LEM），亦被證實會引起小鼠及其他動物之肝癌，因而倍受重視。
   
• 結果判讀
  極限:
  偵測極限(limit of detection, LOD)： 0.3 ppm 
  定量極限(limit of quantification, LOQ)：0.3 ppm
  偵測範圍:0.3–6 ppm(樣品超過6ppm需重新稀釋及測量)
   
• 樣品準備(SAMPLE PREPARATION)
  The sample to be tested should be collected according to accepted sampling techniques (see FGIS sampling protocol or contact your NEOGEN representative). Obtain a representative sample (minimum 100 g). Grind the sample so at least 95% of the ground material passes through a 20 mesh sieve (about the particle size of fine espresso).
   
• 樣品萃取(SAMPLE EXTRACTION)
  1. If not using NEOGEN’s prepared solution, prepare a 65% ethanol solution by mixing 6.5 parts ethanol with 3.5 parts distilled or deionized water for each sample.
  2. Weigh out 10 g ± 0.1 of sample into an extraction cup.
  3. Add 50 mL of 65% ethanol to the extraction cup.
  4. Vigorously shake, using hand or mechanical means, for 3 minutes, or blend for 1 minute.
  5. Allow the sample to settle, then filter with a filter syringe or Whatman #4 filter paper to collect a minimum of 3 mL filtrate into a sample collection tube. Or, you may also pipette 1 mL of sample into a 2.0 mL microcentrifuge tube, and centrifuge for 30 seconds using a microcentrifuge (approx. 2,000 x g).
   
• 樣品萃取(SAMPLE EXTRACTION)— 50 g (FGIS Method)
  1. Combine 50 g of ground sample with 250 mL of 65% ethanol and shake vigorously for 3 minutes or blend for 1 minute. Allow sample to settle for 1–2 minutes.
  2. Filter the extract by pouring 5 mL through a filter syringe and collecting the filtrate. 
  
  1:5 extraction: (1 part sample to 5 parts 65% ethanol), broken rice, canola meal, corn grits, cornmeal, corn, green lentils, oatmeal, soybean meal, wheat flour, wheat.
  1:4 extraction: (1 part sample to 4 parts 65% ethanol), barley, corn gluten meal, DDGS, DDG wet cake, DDG syrup, hominy, milo, oats, rough rice, wheat bran, wheat midds. Please contact NEOGEN for additional extraction information.
  1:3 extraction: (1 part sample to 3 parts 65% ethanol), brewers rice, corn protein concentrate, lentil fiber, navy beans, rice bran, rice hulls, sorghum, sunflower meal. Please contact NEOGEN for additional extraction information.
  NOTE: NEOGEN continues to validate new commodities. Please contact a representative for the latest validated commodity list. 
   
• 操作流程(TEST PROCEDURE)— Raptor Integrated Analysis Platform
  1. Place the appropriate number of red sample dilution cups into a sample cup rack. Label cups if necessary.
  2. Add 200 µL of sample filtrate to each red sample dilution cup.
  3. Add 400 µL of sample diluent to each red sample dilution cup.  Mix by pipetting up and down five times.
  4. Fully insert a Reveal Q+ for Fumonisin test strip into a Raptor cartridge.
  5. Insert the Raptor cartridge containing the test strip into any of the three ports within the Raptor Integrated Analysis Platform reader.
  a. The bar code on the test strip will be read – the Raptor reader identifies the type of test strip and the lot number. If the lot number is not found in the system, the bard code reader on the front of the Raptor will turn on automatically.
  b. Scan the QR code found on the tube containing the test strips. The information will be stored on the system.
  6. Enter a sample ID if desired. Hit the Accept button to go to the Add Sample screen.
  7. Add 400 µL of sample from the red sample dilution cup (from step 3) to the Raptor cartridge.
  a. The Raptor reader will start automatically.
  b. Additional samples can be started in the other ports while the first sample is processing
  8. Results will be displayed on the Raptor screen after the 6-minute testing is complete.
   
• 稀釋步驟(Dilution Procedure) — Raptor Integrated Analysis Platform(樣品超過6ppm需重新稀釋及測量)
  1. Place the appropriate number of red sample dilution cups into a sample cup rack. Label cups if necessary.
  2. Add 200 µL of sample filtrate to a sample collection tube.
  3. Add 200 µL of 65% ethanol. Mix well by pipetting up and down 5 times.
  4. Transfer 200 µL of diluted sample (from step 3) to a new red sample dilution cup.
  5. Add 400 µL of sample diluent (pink-labeled bottle) to the red sample dilution cup. Mix by pipetting up and down five times.
  6. Fully insert a Reveal Q+ for Fumonisin test strip into a Raptor cartridge.
  7. Insert the Raptor cartridge containing the test strip into any of the three ports within the Raptor Integrated Analysis Platform reader.
  a. The bar code on the test strip will be read – the Raptor reader identifies the type of test strip and the lot number.  If the lot number is not found in the system, the bar code reader on the front of the Raptor will turn on automatically.
  b. Scan the QR code found on the tube containing the test strips. The information will be stored on the system.
  8. Enter a sample ID if desired. Hit the Accept button to go to the add sample screen.
  9. Add 400 µL of sample extract from the red sample dilution cup (from step 5) to the Raptor cartridge.
  10. The Raptor reader will start automatically
  11. Additional samples can be started in the other ports while the first sample is processing
  12. Results will be displayed on the Raptor screen after the 6-minute testing is complete. The final result will need to be multiplied by 2. 

• 產品說明書：連結
• 操作手冊：連結
• 物質安全資料表(Safety Data Sheet; SDS)：中文連結、英文連結

參考規範：食品中污染物質及毒素衛生標準