Hereditary spastic paraplegia (HSP) is a neurodegenerative condition defined clinically by lower limb spasticity and weakness. Homozygous mutations in CYP7B1 have been identified in several consanguineous families that represented HSP type 5 (SPG5), one of the many genetic forms of the disease. We used direct sequencing and multiplex ligation-dependent probe amplification to screen for CYP7B1 alterations in apparently sporadic HSP patients (n = 12) as well as index patients from non-consanguineous families with recessive (n = 8) and dominant (n = 8) transmission of HSP. One sporadic patient showing HSP as well as optic atrophy carried a homozygous nonsense mutation. Compound heterozygosity was observed in a recessive family with a clinically pure phenotype. A heterozygous missense change segregated in a small dominant family. We also found a significant association of a known coding polymorphism with cerebellar signs complicating a primary HSP phenotype. Our findings suggest CYP7B1 alterations to represent a rather frequent cause of HSP that should be considered in patients with various clinical presentations.

100 Pre-Contact human skeletons from six different excavation sites on the Peruvian coast, which showed an excellent state of preservation both on the macro-and the microscopic level, have been investigated for the preservation of three plasma proteins (α 2 -HS-glycoprotein, albumin and α 1 -antitrypsin) by enzyme-linked immunosorbent assay. A subsample of 40 skeletons out of this skeletal series had been analysed for the state of preservation of polymorphic plasma proteins by electrophoretic methods (Brandt et al., 2000), revealing extensive alterations of the target proteins by diagenesis. The question remained, therefore, as to the reliability of common immunological tools for the unambiguous detection and quantification of non-collagenous proteins in archaeological bone. In this study, we succeeded in the immunological detection of three target proteins in the skeletal remains. However, a variety of non-specific reactions were also observed with bone extracts from non-human mammals, some collagen types, and also bacterial cells. This wide range of cross-reactions with antigens not normally tested in the course of the production of commercially available antibodies adds a cautionary note to immunological approaches towards the study of ancient proteins.

Protected by the mineral matrix, bone proteins are capable of surviving inhumation periods of several hundreds or thousands of years in soil. While the preservation of the bone matrix protein, collagen I, is the prerequisite for a variety of archaeometric approaches, such as radiocarbon dating and the reconstruction of palaeodiet by stable isotope analysis, little is known about both the rate and state of preservation of non‐collagenous proteins. We succeeded in the isolation, electrophoretic separation (SDS‐PAGE, IEF) and immunological detection (radial immunodiffusion, IEF immunoblotting and ELISA) of plasma proteins preserved in archaeological human bones. However, sample preparation and electrophoretic methods had to be adapted to the specific demands of these aged proteins, since they are not only degraded and fragmented but also cross‐linked to other organic components, either indigenous to the bone or to contaminants from the burial environment. Complex decomposition phenomena are responsible for the altered mode of migration of aged proteins through a gel. After isoelectric focusing, the ancient proteins mainly concentrate below pH 4.45 in the pH‐gradient. Thus, highly negatively charged protein components have a better chance of preservation in bone after death. Isoelectric focusing with subsequent immunoblotting of ancient protein samples revealed protein patterns which showed marked charge‐modifications in comparison with those of modern human plasma proteins due to protein degradation (e.g. α2‐HS‐glycoprotein and α1‐antitrypsin). Nevertheless, in combination with different immunological analyses, previous results concerning the selective enrichment of α2‐HS‐glycoprotein in bone compared with other plasma glycoproteins could be confirmed. Copyright © 1999 John Wiley & Sons, Ltd.

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