ABSTRACT
The National Fish and Wildlife Forensic Laboratory is responsible for the determination of species of birds, reptiles, and mammals from the United States, as well as international species falling under the protection of CITES treaties. We have recently found electrospray ionization mass spectrometry to be an effective means of rapidly analyzing blood samples for species identification. Nearly 1000 individuals were analyzed which comprised 62 species represented by birds, mammals, and reptiles. Whole blood and dried blood samples were analyzed without purification to provide simultaneous molecular weights from the alpha- and beta-proteins present in each sample's hemoglobin. The combination of the two molecular weights for the hemoglobin proteins (i.e., alpha/beta-pairs) was used as species determining markers. In all, 133 distinctive alpha/beta-pairs were observed from the individuals analyzed. Despite the variability in the hemoglobins evaluated, 86% of these alpha/beta-pairs were found to be diagnostic for a particular species to the exclusion of all other species studied. No other single protein system studied by a single analytical technique can so effectively resolve species from a wide range of taxa as can the hemoglobin system when analyzed by electrospray ionization mass spectrometry.
Subject(s)
Animals, Wild/blood , Blood Chemical Analysis/veterinary , Hemoglobins/chemistry , Mass Spectrometry/methods , Animals , Birds/blood , Blood Chemical Analysis/methods , Evaluation Studies as Topic , Mammals/blood , Molecular Weight , Reptiles/blood , Species Specificity , United StatesABSTRACT
The described technique offers a sensitive and reproducible method for inferring the source of over 50 different animal species from bloodstains and blood mixtures. Hemoglobins from each of the species were examined using reversed-phase high performance liquid chromatography (HPLC) in chromatographic times of less than 25 mins. The HPLC method complements and furthers current methodology for identification of species of origin. HPLC analysis is particularly well suited for the quantitative analysis of blood and blood mixtures and is applicable to species for which antisera are unavailable. The sensitivity of the method (hemoglobin amounts down to 1.2 micrograms) lends itself to the analysis of blood mixtures in which only a small percentage of the mixture represents blood from a given species. Such resolution and quantitation is applicable to wildlife forensic casework.
