Stabilization of troponin I for applications in clinical chemistry and forensic medicine. Cardiac troponin I: A time of death marker

Cardiac troponin I (cTnI) is one of the most useful serum marker test for the determination of myocardial infarction (MI). The first commercial assay of cTnI was released for medical use in the United States and Europe in 1995. It is useful in determining if the source of chest pains, whose etiology may be unknown, is cardiac related. Cardiac TnI is released into the bloodstream following myocardial necrosis (cardiac cell death) as a result of an infarct (heart attack). In this research project the utility of cardiac troponin I as a potential marker for the determination of time of death is investigated. The approach of this research is not to investigate cTnI degradation in serum/plasma, but to investigate the proteolytic breakdown of this protein in heart tissue postmortem. If our hypothesis is correct, cTnI might show a distinctive temporal degradation profile after death. This temporal profile may have potential as a time of death marker in forensic medicine. The field of time of death markers has lagged behind the great advances in technology since the late 1850's. Today medical examiners are using rudimentary time of death markers that offer limited reliability in the medico-legal arena. Cardiac TnI must be stabilized in order to avoid further degradation by proteases in the extraction process. Chemically derivatized magnetic microparticles were covalently linked to anti-cTnI monoclonal antibodies. A charge capture approach was also used to eliminate the antibody from the magnetic microparticles given the negative charge on the microparticles. The magnetic microparticles were used to extract cTnI from heart tissue homogenate for further bio-analysis. Cardiac TnI was eluted from the beads with a buffer and analyzed. This technique exploits banding pattern on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) followed by a western blot transfer to polyvinylidene fluoride (PVDF) paper for probing with anti-cTnI monoclonal antibodies. Bovine hearts were used as a model to establish the relationship of time of death and concentration/band-pattern given its homology to human cardiac TnI. The final concept feasibility was tested with human heart samples from cadavers with known time of death. ^

Diatom-based paleolimnological reconstruction of regional climate and local land-use change from a protected sinkhole lake in southern Florida, USA

Despite their sensitivity to climate variability, few of the abundant sinkhole lakes of Florida have been the subject of paleolimnological studies to discern patterns of change in aquatic communities and link them to climate drivers. However, deep sinkhole lakes can contain highly resolved paleolimnological records that can be used to track long-term climate variability and its interaction with effects of land-use change. In order to understand how limnological changes were regulated by regional climate variability and further modified by local land-use change in south Florida, we explored diatom assemblage variability over centennial and semi-decadal time scales in an ~11,000-yr and a ~150-yr sediment core extracted from a 21-m deep sinkhole lake, Lake Annie, on the protected property of Archbold Biological Station. We linked variance in diatom assemblage structure to changes in water total phosphorus, color, and pH using diatom-based transfer functions. Reconstructions suggest the sinkhole depression contained a small, acidic, oligotrophic pond ~11000–7000 cal yr BP that gradually deepened to form a humic lake by ~4000 cal yr BP, coinciding with the onset of modern precipitation regimes and the stabilization of sea-level indicated by corresponding palynological records. The lake then contained stable, acidophilous planktonic and benthic algal communities for several thousand years. In the early AD 1900s, that community shifted to one diagnostic of an even lower pH (~5.6), likely resulting from acid precipitation. Further transitions over the past 25 yr reflect recovery from acidification and intensified sensitivity to climate variability caused by enhanced watershed runoff from small drainage ditches dug during the mid-twentieth Century on the surrounding property.

The status and ecology of the American alligator (Alligator mississippiensis) in Par Pond, Savannah River site

Par Pond is a man-made 1120 ha cooling reservoir located on the Savannah River Site near Aiken, South Carolina. From 1972-1978 a detailed study on the status of the alligator in Par Pond was conducted by Tom Murphy (unpub. MS thesis Univ. of GA, 1977). Murphy estimated that approximately 110 alligators inhabited Par Pond with an adult (> 1.8 m) to juvenile (< 1.8 m) ratio of (1.8:1), an overall sex ratio of 3.2:1, and an average of only 2.3 nests/yr. The purpose of this study (1986-1989) was to determine the current population size and structure, determine how the population has changed in the last 15 years and to examine growth and survival of juvenile alligators. Data were collected by monthly night-time eyeshine counts aerial surveys, capturing animals, and locating and following the fate of nests. There was a strong positive correlation between water temperature and the number of alligators observed during eyeshine counts. Both eyeshine counts and aerial surveys were highest in spring and varied seasonally. A total of 184 different non-hatchling and 157 hatchling alligators were captured between May 1986 and November 1988. Population estimates and size distributions based on capture data indicate that over the last 15 years the population has increased from approximately 110 to 200 alligators, and the size distribution has shifted from one dominated by large adults to one that has a higher proportion of juveniles. The current sex ratio (2.6:1) is not significantly different from that reported by Murphy (1977, 3.2:1). However, the average number of nests/yr has increased from 2.3 to 4.0. Data on juvenile growth and survival show that the growth rate of hatchlings (32.9 cm/yr total length) is greater than that of animals age 1-3 (21.6 cm/yr total length) and survival of all ages is variable between years and between clutches. Results from this study indicate that from 1972-988 the population has increased ac an average exponential rate of 6 % per year. If conditions in Par Pond do not change, the population size should continue to increase.