1 / 19

EURO-DIAGNOSTICA B.V. The Netherlands For all information, please contact authorized European representative and distr

EURO-DIAGNOSTICA. EURO-DIAGNOSTICA B.V. The Netherlands For all information, please contact authorized European representative and distributor: Diagnostics Center “EUROTEST” d.o.o. Sarajevo, Bosnia and Herzegovina, Europe Tel/fax: +387 (0)33-555-801 , Cell Phone: +387 (0) 61 - 201 - 711

apria
Download Presentation

EURO-DIAGNOSTICA B.V. The Netherlands For all information, please contact authorized European representative and distr

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. EURO-DIAGNOSTICA EURO-DIAGNOSTICA B.V. The Netherlands For all information, please contact authorized European representative and distributor: Diagnostics Center “EUROTEST” d.o.o. Sarajevo, Bosnia and Herzegovina, Europe Tel/fax: +387(0)33-555-801, Cell Phone: +387(0)61-201-711 E-mail: info@eurotest.co.ba www.eurotest.co.ba

  2. EIA TECHNOLOGY FOR THE DETECTION OF THE ANTIBIOTIC CHLORAMPHENICOL (CAP) IN SHRIMPS

  3. EIA: ENZYME IMMUNOASSAY EIA is a rapid test used for detecting and quantifying a large variety of biological components, e.g. viruses, bacteria, hormones, antibiotics, allergens, etc.

  4. EIAs are divided into 3 main formats: • Direct EIAs • Indirect EIAs • Blocking or competitive EIAs EIA FORMATS The CAP-EIA is a competitive assay!

  5. CAP-EIA KIT FROM EURO-DIAGNOSTICA

  6. REAGENTS IN THE CAP-EIA • A 96-wells polystyrene microtitre plate • coated with sheep anti-rabbit antibodies • Rabbit anti-CAP antibodies • Conjugate: labeled CAP (CAP-PO) • CAP-standards, negative controls • Sample material (not in the kit!)

  7. MICROTITRE-PLATE IN THE CAP-EIA The wells of the microtitre plate are coated with sheep anti-rabbit (SAR) antibodies. The function of these antibodies is to capture the rabbit anti-CAP antibodies.

  8. IMMUNISATION OF RABBIT CAP COUPLING OF CAPTO A CARRIER PROTEIN ANTIBODIES ARE PURIFIED AND USED IN THE CAP-EIA RABBIT PRODUCES ANTI-CAP ANTIBODIES ANTI-CAP ANTIBODIES IN THE EIA HAVE BEEN RAISED IN RABBITS

  9. + CAP PO CAP-PO (CONJUGATE) CONJUGATE IN THE CAP-EIA THE CONJUGATE IN THE CAP-EIA CONSISTS OF CAP THAT HAS BEEN LABELED WITH THE ENZYME HORSE-RADISH PEROXIDASE (PO)

  10. SHRIMPS CONTAINING CAP HOMOGENISATION OF SHRIMPS IN BLENDER EXTRACTION OF CAP WITH ETHYL ACETATE RECONSTITUTION IN DILUTION BUFFER SAMPLE IN CAP-EIA SAMPLE PREPARATION CAP-EIA

  11. RABBIT ANTI CAP ANTIBODIES C CONJUGATE B STANDARDS, SAMPLES, NEG. CONTROLS A SHEEP ANTI-RABBIT ANTIBODIES REACTION WELL STEPS IN THE CAP-EIA 1. THE SEVERAL REAGENTS ARE ADDED IN ORDER A, B, C

  12. STEPS IN THE CAP-EIA 2. THE PLATE IS INCUBATED FOR 60 MIN AT 4˚C 3. THE WELLS ARE THEN WASHED WITH WASHING BUFFER • TWO SITUATIONS CAN NOW BE CONSIDERED: •  THE SAMPLE CONTAINS A CERTAIN AMOUNT OF CAP •  THE SAMPLE CONTAINS NO CAP

  13. STEPS IN THE CAP-EIA SAMPLE WITH CAP: SAMPLE WITHOUT CAP: CONJUGATE AS WELL ASFREE CAP CAN BIND TO THE ANTI-CAP ANTIBODIES ONLY THE CONJUGATE IS AVAILABLE TO BIND TO THE ANTI-CAP ANTIBODIES

  14. STEPS IN THE CAP-EIA 4. THE ENZYME SUBSTRATE/CHROMOGEN (H2O2/TMB) IS ADDED DURING INCUBATION THE COLORLESS CHROMOGEN IS CONVERTED BY THE ENZYME INTO A BLUE REACTION PRODUCT. TMB/H2O2 5. THE PLATE IS INCUBATED FOR 30 MIN AT 25˚C

  15. STEPS IN THE CAP-EIA THE BLUE COLOR IS INVERSELY PROPORTIONAL TO THE AMOUNT OF BOUND CAP. THE MORE CAP PRESENT IN THE SAMPLE, THE LESS COLOR IS DEVELOPED. H2O2/TMB

  16. STEPS IN THE CAP-EIA 6. THE COLOR DEVELOPMENT IS STOPPED BY ADDING 0.5 M OF SULFURIC ACID (H2SO4) IN THIS ACIDIC ENVIRONMENT THE BLUE COLOR CHANGES INTO YELLOW

  17. READING THE RESULTS 7. ABSORPTION OF EACH WELL IS MEASURED AT 450 nm 8. AMOUNT OF CAP IN THE SAMPLES IS CALCULATED

  18. Greetings from Holland Thanks for your attention!

  19. CONTACT EURO-DIAGNOSTICA EURO-DIAGNOSTICA B.V. The Netherlands For all information, please contact authorized European representative and distributor: Diagnostics Center “EUROTEST” d.o.o. Sarajevo, Bosnia and Herzegovina, Europe Tel/fax: +387(0)33-555-801, Cell Phone: +387(0)61-201-711 E-mail: info@eurotest.co.ba www.eurotest.co.ba

More Related