V. Cortellini

Application of alternative fixatives to formalin in diagnostic pathology.

Fixation is a critical step in the preparation of tissues for histopathology. The objective of this study was to investigate the effects of different fixatives vs formalin on proteins and DNA, and to evaluate alternative fixation for morphological diagnosis and nucleic acid preservation for molecular methods. Forty tissues were fixed for 24 h with six different fixatives: the gold standard fixative formalin, the historical fixatives Bouin and Hollande, and the alternative fixatives Greenfix, UPM and CyMol. Tissues were stained (Haematoxylin-Eosin, Periodic Acid Schiff, Trichromic, Alcian-blue, High Iron Diamine), and their antigenicity was determined by immunohistochemistry (performed with PAN-CK, CD31, Ki-67, S100, CD68, AML antibodies). DNA extraction, KRAS sequencing, FISH for CEP-17, and flow cytometry analysis of nuclear DNA content were applied. For cell morphology the alternative fixatives (Greenfix, UPM, CyMol) were equivalent to formalin. As expected, Hollande proved the best fixative for morphology. The morphology obtained with Bouin was comparable to that with formalin. Hollande was the best fixative for histochemistry. Bouin proved equivalent to formalin. The alternative fixatives were equivalent to formalin, although with greater variability in haematoxylin-eosin staining. It proved possible to obtain immunohistochemical staining largely equivalent to that following formalin-fixation with the following fixatives: Greenfix, Hollande, UPM and CyMol. The tissues fixed in Bouin did not provide results comparable to those obtained with formalin. The DNA extracted from samples fixed with alternative fixatives was found to be suitable for molecular analysis.

Population data on 5 non-CODIS STR loci (D10S1248, D22S1045, D2S441, D1S1656, D12S391) in a population sample from Brescia county (Northern Italy)

Abstract Five autosomal STR loci (D10S1248, D22S1045, D2S441, D1S1656 and D12S391), unlinked to the core STR loci (non-CODIS loci), were typed in a population sample of 210 unrelated individuals from Brescia (Northern Italy). Allelic frequencies and statistical parameters of forensic interest were estimated. A comparison between our population data and others from Caucasian and Asian populations was performed.

Post-mortem injuries by a dog: a case report

Despite its unlikely occurrence, post-mortem animal depredation is not unknown to forensic pathologists. In the case at issue, the corpse of a dead woman presented extensive facial wounds, which were then traced back to the dog she owned. A small specimen of injured tissue was subjected to species diagnosis, and came back positive for human and canine antigens, which confirmed the presence of biological material of canine origin on the body. The less than usual post-mortal injury pattern described herein clearly highlights the possibility that animal depredation on a corpse may occur soon after death, and underscores the diagnostic potential posed by commonly available and low expensive testing methods such as serological species diagnosis.

Y-chromosome polymorphisms and ethnic group – a combined STR and SNP approach in a population sample from northern Italy

Aim To find an association between Y chromosome polymorphisms and some ethnic groups. Methods Short tandem repeats (STR) and single-nucleotide polymorphisms (SNP) on the Y chromosome were typed in 311 unrelated men from four different ethnic groups – Italians from northern Italy, Albanians, Africans from the Maghreb region, and Indo-Pakistanis, using the AmpFlSTR® Yfiler PCR Amplification Kit and the SNaPshot Multiplex Kit. Results STRs analysis found 299 different haplotypes and SNPs analysis 11 different haplogroups. Haplotypes and haplogroups were analyzed and compared between different ethnic groups. Significant differences were found among all the population groups, except between Italians and Indo-Pakistanis and between Albanians and Indo-Pakistanis. Conclusions Typing both STRs and SNPs on the Y chromosome could become useful in determining ethnic origin of a potential suspect.