Monoclonal Antibodies as Probes for the Detection of Porcine Blood-Derived Food Ingredients

Abstract

The lack of effective methods to monitor the use of porcine blood-derived food ingredients (PBFIs) is a concern for the billions of individuals who avoid consuming blood. We therefore sought to develop a panel of porcine blood-specific monoclonal antibodies (mAbs) for use as probes in immunoassays for the detection of PBFIs. Ten selected mAbs were identified that react with either a 60 or 90 kDa protein in the plasma fraction or a 12 kDa protein in the red blood cell fraction of porcine blood. Western blot analysis of commercially produced PBFIs revealed that these antigenic proteins are not affected by various manufacturing processes. The utility of these mAbs was demonstrated in a prototype sandwich ELISA developed for this study using mAbs 19C5-E10 and 16F9-C11. The new assay is porcine blood-specific and capable of detecting ≤0.03% (v/v) of PBFIs in cooked (100 °C for 15 min) ground meats or fish.

Development of Monoclonal Antibodies Recognizing Linear Epitope: Illustration by Three Bacillus thuringiensis Crystal Proteins of Genetically Modified Cotton, Maize, and Tobacco

“…The larger molecular protein on lane 5 could be the cotton Cry1Ac protein precursor. The small molecular protein could be either the degraded peptides during extraction or another small protein bearing the epitope that could be recognized by the antibody . Western blot assay could be used to study tissue-specific protein expression due to different post-translation modification .…”

“…The small molecular protein could be either the degraded peptides during extraction or another small protein bearing the epitope that could be recognized by the antibody. 41 Western blot assay could be used to study tissuespecific protein expression due to different post-translation modification. 42 The result of the present study indicated that Western blot could give more information on protein heterogeneity than ELISAs because Western blot employs the electrophoresis separation power and antibody's high specificity and affinity.…”

Non-Specific Binding and Cross-Reaction of ELISA: A Case Study of Porcine Hemoglobin Detection

“…An enzyme inhibitor, Halt Protease Inhibitor Cocktail (78425), was purchased from Thermo Scientific (Rockford, IL, USA). Mouse anti-P Hb mAb was developed at the Florida State University Hybridoma Facility (Tallahassee, FL, USA) [26], and rabbit anti-P Hb pAb was developed at the Hebei Animal Disease Prevention and Control Center (Shijiazhuang, Hebei, China) [24]. Mouse anti-P Hb mAb was biotinylated using EZ-link sulfo-NHS-LC-Biotin (sulfosuccinimidyl-6-[biotin-amido]hexanoate, 21335, Thermo Scientific) according to the manufacturer's instructions.…”

Progress and challenges associated with halal authentication of consumer packaged goods

“…A detection level of up to 1% was achieved for meat bone meal in soy products without the need for complicated sample processing steps . Recently, a sandwich enzyme‐linked immunosorbent assay (ELISA) utilizing a monoclonal antibody was developed for food ingredients derived from porcine blood with a detection limit as low as 0.03% (v/v) . A rapid lateral flow immuno‐chromatographic assay utilizing thermostable meat protein for the detection of raw, cooked meat and gelatin samples was developed that was capable of detecting as low as 0.01% based on the sample matrix, with no cross‐reactivity over related species .…”