Biochemical analytes, sample matrices, and potential forensic utility.
Abstract
The diagnosis of acute cardiac pathology is a clinical challenge in both the living and in the postmortem setting. Cardiac troponin (cTn) T and cardiac troponin I released from the contractile apparatus of cardiomyocytes into the circulation can be detected by sensitive and specific immunoassays and are the gold standard biochemical test for diagnosis of acute coronary syndromes (ACS). Recently with the advent of more sensitive detection methods, elevation in non-ACS has become apparent causing clinical confusion. In most cases, these elevations are related to subclinical cardiac damage and often confer poor prognosis in cTn-positive patients. Biomarkers of cardiomyocyte damage may be of value in routine hospital and medico-legal autopsy. A significant body of evidence has emerged since the late 1990s, assessing the clinical utility of cardiac troponin in biological fluids or in immunohistochemical staining of cardiac tissue to aid in the diagnosis of acute cardiac pathology when standard microscopic evidence is inconclusive. This chapter reviews the extensive literature on the subject and details the disparity between pericardial fluid and serum for the use of cTn in the postmortem setting.
Keywords
- cardiovascular disease
- risk
- diagnostics
- therapeutic intervention
- treatment
- prediction
1. Introduction
Cardiac troponins (cTn) T (cTnT) and I (cTnI) are the gold standard biochemical markers used to identify acute cardiac pathologies in patients who present with typical and atypical chest pain to the emergency department. These muscle-associated proteins confer superior diagnostic and prognostic ability compared to conventional nonspecific muscle derived enzyme markers such as creatine kinase (CK), its MB isoform (CK-MB), or myoglobin. Cardiac troponin determination is central to the diagnosis of non-ST segment elevation acute myocardial infarction (NSTEMI), contributing to international guidelines for diagnosis and management of NSTEMI patients [1].
Recent advancement in laboratory technology driven both by clinical demand and the commercial
One area of interest has been the potential value of cTnT and cTnI in the postmortem setting and may provide insight into the cause of death. Troponin analysis in postmortem blood and pericardial fluid during autopsy investigations can potentially help medical examiners and forensic pathologists attribute what happened before, during, and after a death. This chapter will explore the use of cardiac troponin in the postmortem setting, from its application in routine hospital as well as medico-legal autopsy and forensics, assessing the usefulness in offering a clearer picture of an individual’s final moments.
2. Clinical utility of cardiac troponin in myocardial damage
Cardiac-specific isoforms of the contractile protein complex troponin, namely cTnT and cTnI, are released into the bloodstream following damage to cardiomyocytes. The mechanism by which these structural proteins are released into the circulation has been debated significantly over many years. Initially, it was thought that cTn could only be released following overt cellular necrosis; however, recently it has been suggested that release can occur in ischemia without necrosis [3]. A review of the subject by Ragusa and colleagues suggest release mechanisms, including apoptosis, necroptosis, physiological cardiomyocyte renewal, and cellular wounding can contribute to cTn release as well as necrosis [4]. An immunohistochemical study using a canine model of coronary occlusion ranging from 30 minutes to 6 hours demonstrated variable loss of both cTnT and cTnI in paraffin-embedded left ventricular myocardial sections [5]. Loss was variable but more so for cTnT than for cTnI, and loss was greater at the periphery of the infarct area rather than the centralised region (Figure 1).
Using monoclonal antibodies specific to the cardiac isoforms, immunoassay technologies can quantify the amount of cTnT or cTnI in a biological matrix [6]. Initially, early immunoassays utilised high clinical cut-off values (high specificity and low sensitivity) allowed the separation of patients with overt acute myocardial infarction (AMI) from apparently healthy persons who were deemed negative for cTn based on the equivalent cTnT or cTnI concentration to the then-used gold standard tests (CK or CK-MB). Subsequently, the large body of evidence demonstrating elevation of CK and CK-MB in the absence of an elevated cTn questioned the cardio-specificity of the enzyme markers, along with approximately 30% of patients ruled out with AMI demonstrating positive cTn which is associated with poor prognosis, resulted in the adoption of cTn as the gold standard test for diagnosis of AMI [6].
Integral to the adoption of cTn was the appropriate definition of cut-off to confer an abnormal concentration. This was subsequently defined as the 99th percentile value of an apparently healthy population. When adopted into routine clinical practice, this lowered the sensitivity of the assays allowing early diagnosis in the evolving infarction but at the cost of specificity. Initially, this caused clinical confusion with a larger number of patients presenting with low concentrations of cTn just above the AMI cut-off value, but further research of such patients found the presence of comorbid conditions (Figure 2) often with underlying cardiovascular pathophysiology [2].
3. Biochemical testing in assisting cause of death at postmortem
Biochemical testing in postmortem investigations (termed thanatochemistry, necrochemistry or the chemistry of death) was initially established in the early 1950s, and a great number of biochemical analytes have proved an adjunctive tool to assist the cause of death at postmortem [7]. Adoption of biochemical testing especially in the medico-legal forensic autopsy has often been limited. The determination of death may have significant impact on those directly or indirectly involved in the death of an individual and can carry a custodial sentence. Thus, the scientific evidence presented in court is intensely scrutinised both by the prosecution and defence counsels. Whilst no biochemical test is infallible, many are associated with the likelihood of a disease process rather than a definitive diagnosis of the disease. Often the barrier to use is the interpretation of results of biochemical assays from cadaveric sampling, hindered by the lack of reference normality in death; thus, results are compared to reference intervals generated in the living [7, 8] with few studies demonstrating corresponding histopathological findings to the biochemical results. Interpretation is further complicated by factors such as postmortem interference in the assay technology, appropriate sampling matrices, postmortem autolysis, microbial metabolism, fluid redistribution, and postmortem interval (PMI). Molecular biophysical properties such as molecular weight, structure, intracellular location, electrical charge, ionic strength, protein affinity, and cell membrane permeability may differ between life and death and can influence interpretation in both situations [7].
There are a number of fluid components which are suitable for cadaveric biochemical testing, namely vitreous humour from the posterior segment of the eye, cerebral spinal fluid (CSF), synovial fluid, pericardial fluid (PCF), venous femoral blood, venous jugular blood, peripheral blood sampling, urine, gastric contents and right ventricle heart whole blood [7, 8, 9]. Analytes and potential uses in postmortem samples are listed in Table 1.
Analyte | Sample matrix | Forensic utility |
---|---|---|
Adrenaline:Noradrenaline | Urine | Hypothermia |
Acetone | Blood | Chronic alcohol abuse, hypothermia, diabetic ketoacidosis |
Ammonia | Vitreous humour | Liver failure |
Carbohydrate-deficient transferrin | Vitreous humour | Chronic alcohol abuse |
Chloride | Vitreous humour | Saline poisoning, salt water drowning, dehydration |
Chymase activity | Blood | Anaphylactic shock |
Chromogranin A | Blood, CSF | Hypothermia |
C-reactive protein | Blood | Recent infection, trauma, burns, ketoacidosis, malignancy, autoimmune diseases, inflammatory diseases, sepsis |
Creatine Kinase & CK-MB | Blood | Cardiac pathology |
Creatine Kinase-BB | CSF | Cerebral trauma, cerebral hypoxia |
Creatinine | Vitreous humour | AKI, CKD, high-protein diet, large muscle mass (anabolic steroid abuse), heat shock |
Ethyl glucuronide | Vitreous humour, Urine | Antemortem ethanol ingestion |
Free fatty acids | Blood | Hypothermia |
Fructoasmine | Vitreous humour | Diabetic ketoacidosis |
Glucocortcoids | Blood | Hypothermia |
Hypoxanthine | Vitreous humour | Time of death |
Myoglobin | Blood, Urine | Hyperthermia, cardiac pathology |
Neurone specific enolase | CSF | Cerebral traumatic injury, cerebral hypoxia |
Potassium | Vitreous humour | Postmortem decomposition |
S-100b | CSF | Cerebral injury |
Thyroglobulin/fT3 | Vitreous humour, Blood | Neck trauma, strangulation |
Troponin | Pericardial fluid, Blood | Cardiac pathology |
Tryptase | Blood | Anaphylactic shock |
Urea | Vitreous humour | Renal dysfunction, upper GI haemorrhage |
4. Conventional cardiac biomarkers at postmortem
The importance of cardiac biomarkers assisting in postmortem diagnosis was highlighted in cases where a suspected myocardial lesion cannot be diagnosed by routine histological analysis. They were utilised initially for the determination of sudden cardiac death. Initially, CK and lactate dehydrogenase isoenzyme analysis of pericardial fluid was utilised [10, 11], followed by K:Na ratio [12]; CK isoenzymes, aspartate aminotransferase and hydroxybutyrate dehydrogenase [13] and myosin and cathepsin D, a lysosomal aspartyl protease that degrades proteins [14, 15].
5. Cardiac troponin analysis at postmortem
The first reported use of cTn analysis was in 1998 by Osuna and colleagues who studied 89 cadavers with a mean age of 51.38 ± 2.04y [16]. Subjects were assigned between four groups, MI (n = 25), asphyxia (n = 30), cranio/multiple trauma (n = 17), and other natural deaths (n = 17). MI was determined by H&E and acridine orange histological staining. The research group determined the concentration of myoglobin, myosin, CK-MB, and cTnI in femoral vein serum samples and PCF in each case. PCF concentrations for all makers were significantly different between each outcome group; however, only myoglobin and myosin demonstrated significance in serum. The PCF cTnI values were higher than serum samples when using the Sanofi Diagnostic Pasteur assay. Values in both matrices were higher in MI patients compared to the other three groups (mean (range) Pericardial cTnI [pg/L]): 2.4 (0.3–6.5); 1.7 (0.03–3.7); 1.1 (0.01–2.3); 0.4 (0.0–1.8) in each group, respectively.
Cina and colleagues [17] demonstrated the utility of cTnT using the then available commercial Cardiac Rapid T (cTnT) lateral flow test from Roche Diagnostics. This device allows testing in the autopsy suite at the time of postmortem with qualitative results (positive or negative test lines) at 15 minutes from sample application. In 40 autopsy cases, 20 were deemed cardiac deaths and 20 were controls (noncardiac related) deaths, diagnosed by gross pathology and histological analysis. 85% (n = 17) of subclavian or femoral blood samples in the cardiac death group were positive for cTnT which was significantly different to control group, where 30% (n = 6) of serum samples were positive for cTnT. The authors deemed these as false-positive results. In subjects aged over 50 years, sensitivity and specificity of cTnT for diagnosis of AMI were 91% and 86%, respectively. The authors noted however the assay was ineffective in frozen blood samples or those with significant haemolysis which was evident at a PMI of >24 h. A similar study of 100 autopsy cases of sudden unexplained death (SUD) was carried out in Chaing Mai, Thailand, and utilised the same rapid cTnT assay [18]. Fifty-two of the deaths were considered cardiac with 20 due to sudden MI and 32 with evidence of old infarction or arrhythmic fibrosis (n = 22), coronary atherosclerosis, >75% luminal without evidence of fibrosis or thrombosis (n = 3), cardiomegaly, and heart weight > 400 g (n = 7) or related to cardiac injury as a result of toxic substances. Thus, subjects were assigned to other cardiac death (SCD), non-cardiac natural death (NCD) or non-natural death (NND). The percentage positivity rate was higher in subclavian blood than femoral blood samples in all three groups. Subclavian blood sensitivity and specificity for SUD were 87.5% and 47%, respectively. Similar to the findings of Cina and colleagues, false-positive rates were associated with increasing PMI.
Davies and colleagues were the first to compare antemortem (<48 h before death) and postmortem concentrations of cTnT (Roche Elecsys 3rd generation electrochemiluminescent assay) and cTnI (Stratus CS fluorometric assay, Dade-Behring [now Siemens healthineers]) in five hospital-based autopsies [19]. One patient suffered cardiac death (myocarditis) with the remaining four were non-cardiac, but moribund before death. Results obtained between antemortem and postmortem samples were erratic. Four of the five subjects (n = 80%) had elevated antemortem cTnT and cTnI samples. The authors concluded postmortem cTn analysis in blood was not suitable due to lack of correlation of cause of death; however, they suggested that elevated antemortem cTn was related to all-cause mortality in those at end of life. A similar conclusion was made by Rahimi and colleagues in 2018 after studying 140 natural and unnatural deaths in Malaysia [20]. Subjects were classified into five groups: cardiovascular death, sudden unexplained death, thoracic trauma, non-thoracic trauma, and other diseases. Median jugular/subclavian/femoral blood cTnT (Roche Elecsys 3rd generation electrochemiluminescent assay) concentrations were 0.51, 0.17, 0.62, 0.90, and 0.51 μg/L, respectively, with no significant difference (p= > 0.05) in relation to cause of death. The authors concluded cTnT lacked specificity in postmortem sampling and is therefore not a useful tool.
Lai and colleagues also compared antemortem and postmortem blood sampling. Demonstrating in four cases, a marked proportionate rise in cTnT in postmortem samples compared to antemortem samples. Interestingly, the authors found cTnT values were higher (mean cTnT 5.32 μg/L) in non-cardiac deaths compared to 4.91 μg/L in cardiac-related deaths [21].
In 2006, Zhu and co-workers published two seminal papers in
Remmer and colleagues [24] focused on postmortem serum and PCF cTnT in relation to PMI from 101 forensic autopsy cases in Estonia. PMI ranged from 8 h to 141 h. Although differences in cTnT were observed between five groups of cause of death (cardiovascular disease; other disease; poisoning; asphyxia; drowning; hypothermia; thoracic trauma, other trauma and fatal fires), significant attention to PMI (Figure 3) is important rather than comorbid cardiovascular disease.
In addition to the effects of PMI as demonstrated above, Kumar et al. used SDS-PAGE and Western blotting of cardiac tissue extracts from 10 medicolegal autopsies of burns cases [25]. The authors demonstrated a pattern of cTnT degradation in a time-dependent manner at room temperature (7.3 h, 18.2 h, 30.3 h, 41.2 h, 41.4 h, 54.3 h, 65.2 h and 88.4 h), demonstrating the disappearance of intact cTnT protein and the increasing presence of low-molecular-weight bands related to time (Figure 4a). Furthermore, the groups have examined degradation patterns according to the cause of death (Figure 4b–d) [26].
A study of 20 autopsies of sudden cardiac death and 8 controls (violent non-cardiac deaths) demonstrated significantly higher cTnT and cTnI concentrations in pulmonary venous blood. Mean ± SD cTnT were 1826 ± 363 μg/L versus 65 ± 11 μg/L, respectively, and for cTnI were 28 ± 3 mg/L versus 0.14 ± 0.02 μg/L; however, it should be noted that the PMI in all cases was 8 h [27].
The value of cTn as a biochemical marker in relation to sudden cardiac death has been the subject of a systematic review [28] and formal meta-analysis [29]. Whilst both reviews demonstrate the elevation of cTn to be higher in pericardial fluid compared to blood sampling in the postmortem setting (Figure 5), blood is susceptible to the effects of haemolysis, postmortem interval, autolysis, and potential bacterial interferences. Pericardial fluid is therefore the preferred sample of choice.
Both reviews also address the issue of cut-off values demonstrating significant difference to cut-off values in the living. Non-cardiac deaths often demonstrate significantly positive cTn values in PCF and blood, thus questioning the sensitivity and specificity at postmortem. Barberi and van den Hondel suggest that more work is required to determine the appropriate cut-off values at postmortem [28].
6. Correlation between postmortem cardiac troponin and histological evidence of cardiomyocyte necrosis
At postmortem, the diagnosis of myocardial infarction is typically assessed by gross macroscopic anatomy further confirmed by microscopic histology and immunohistochemical analysis. Defining AMI in a medico-legal autopsy is a clinical challenge for the forensic pathologist as detection can only be made 4–6 hours after the onset of cardiac ischemia. Histological changes indicative of AMI include oedema, congestion, haemorrhage, inflammation cytoplasmic vacuoles, contraction band alterations, fibrosis and necrosis. Immunohistochemical analysis at postmortem has focused on a number of markers of cellular damage, including C5b-9, myoglobin, CK-MB, fibronectin myosin heart-type fatty acid binding protein and desmin [30].
A number of studies have evaluated postmortem cTn concentrations in relation to evidence of cardiomyocyte necrosis (Figure 6). Ortmann and colleagues identified antigen depletion in the detection of early ischemic cardiac lesions in 8 cases of AMI, 8 cases of sudden cardiac death and 12 cases of acute exogenic hypoxia due to hanging or carbon monoxide poisoning. Strong evidence of immunohistochemical depletion of cTnT was evident in all eight cases of AMI, in 50% of sudden cardiac death and in 1 (8%) of acute exogenic hypoxia, with 42% demonstrating weak loss and 50% demonstrating negative results (no loss of cTnT staining) [30].
Martinez Diaz and colleagues have demonstrated immunohistological changes in cTn in AMI or multiple trauma compared to other causes of death. PCF cTnI, myoglobin and CKMB were all significantly higher in AMI or multiple trauma cases compared to other causes of death. Serum concentrations of cTnI myoglobin and CKMB were not significantly different between the two groups [31]. Immunohistological analysis was performed by the authors with the analysis of troponin C and cTnT staining. 86% of cases demonstrated strong positive TnC with expression differing in isolated cells demonstrating contraction band necrosis but with significantly less intensity in the area of the infarction. cTnT staining was less evident in only 46% of cases in focal areas of the tissue.
Campobasso et al. [34] compared 4 immunohistochemical markers as early indicators of myocardial ischemia in 18 sudden cardiac deaths (4 AMI,4 coronary deaths, 8 acute cardiac deaths compared to 6 cases of acute traumatic death gunshot wounds with immediate lethal head injury). The authors stained paraffin-embedded myocardial tissue and immunohistochemically stained the tissue for C5b-9, fibronectin, myoglobin and cTnI. Diffuse depletion of cTnI was evident in all AMI deaths, in 75% of acute cardiac deaths, with 50% of coronary deaths demonstrating limited cellular foci depletion and normal distribution in all six cases of acute traumatic death. Whilst the staining patterns were significantly different between the cardiac and non-cardiac deaths, the authors concluded that no single marker was able to detect early myocardial ischemia and the combination of all four markers was useful in demonstrating evidence of myocardial ischemia and/or necrosis [34].
More recently, Amin and co-workers stained histological sections from ischemic and non-ischemic cardiac tissue for cTnT, myoglobin and caspase-3, demonstrating cTnT detection in normal myocardium and loss in necrotic tissue. The loss of cTnT was non-uniform with greater loss at the periphery compared to the central regions of infarcted tissue [35].
7. Conclusions
This chapter summarises the extensive literature base examining the clinical utility of cardiac troponin when tested in the postmortem setting. Whilst there is overwhelming evidence to support the superior value of pericardial fluid cTn rather than blood sampling due to significant interferences with the latter, there remains the issue of clinically validated cut-off concentrations in postmortem sampling. The effect of autolysis and increasing concentrations of cTn in fluid analysis correlated with increasing postmortem interval of significance. These features therefore suggest that cTn analysis is more suited as a rule out of cardiac involvement in sudden death rather than a rule in diagnostic aid. The diagnostic utility should be limited to the hospital autopsy rather than the medico-legal postmortem where these factors and interferences could provide scope for counter arguments by the defence counsel. Further work is required in the medico-legal setting to establish appropriate diagnostic cut-off values for cTnT and cTnI in postmortem samples, and clinical pathological guidelines should be written to provide support to the forensic teams to correctly interpret the evidence from this large selection of published literature.
References
- 1.
Thygesen K et al. Fourth universal definition of myocardial infarction (2018). European Heart Journal. 2019; 40 (3):237-269 - 2.
Park KC et al. Cardiac troponins: From myocardial infarction to chronic disease. Cardiovascular Research. 2017; 113 (14):1708-1718 - 3.
Hickman PE et al. Cardiac troponin may be released by ischemia alone, without necrosis. Clinica Chimica Acta; International Journal of Clinical Chemistry. 2010; 411 (5-6):318-323 - 4.
Ragusa R et al. Cardiac troponins: Mechanisms of release and role in healthy and diseased subjects. BioFactors (Oxford, England). 2023; 49 :251-264 - 5.
Fishbein MC et al. Myocardial tissue troponins T and I. An immunohistochemical study in experimental models of myocardial ischemia. Cardiovascular Pathology: The Official Journal of the Society for Cardiovascular Pathology. 2003; 12 (2):65-71 - 6.
Collinson PO et al. Measurement of cardiac troponins. Annals of Clinical Biochemistry. 2001; 38 (Pt 5):423-449 - 7.
Luna A. Is postmortem biochemistry really useful? Why is it not widely used in forensic pathology? Legal Medicine (Tokyo, Japan). 2009; 11 (Suppl 1):27 - 8.
Belsey SL, Flanagan RJ. Postmortem biochemistry: Current applications. Journal of Forensic and Legal Medicine. 2016; 41 :49-57 - 9.
Madea B, Musshoff F. Postmortem biochemistry. Forensic Science International. 2007; 165 (2-3):165-171 - 10.
Luna A et al. The determination of CK, LDH and its isoenzymes in pericardial fluid and its application to the post-mortem diagnosis of myocardial infarction. Forensic Science International. 1982; 19 (1):85-91 - 11.
Stewart RV et al. Postmortem diagnosis of myocardial disease by enzyme analysis of pericardial fluid. American Journal of Clinical Pathology. 1984; 82 (4):411-417. [Accessed: February 22, 2023] - 12.
Lachica E et al. Comparison of different techniques for the postmortem diagnosis of myocardial infarction. Forensic Science International. 1988; 38 (1):21-26 - 13.
Burns J et al. Necropsy study of association between sudden death and cardiac enzymes. Journal of Clinical Pathology. 1992; 45 (3):217-220 - 14.
Perez-Cárceles MD et al. Biochemical assessment of acute myocardial ischaemia. Journal of Clinical Pathology. 1995; 48 (2):124-128. [Accessed: February 22, 2023] - 15.
Pérez-Cárceles MD et al. Usefulness of myosin in the postmortem diagnosis of myocardial damage. International Journal of Legal Medicine. 1995; 108 (1):14-18. [Accessed: February 22, 2023] - 16.
Osuna E et al. Cardiac troponin I (cTn I) and the postmortem diagnosis of myocardial infarction. International Journal of Legal Medicine. 1998; 111 (4):173-176 - 17.
Cina SJ et al. A rapid postmortem cardiac troponin T assay: Laboratory evidence of sudden cardiac death. The American Journal of Forensic Medicine and Pathology. 2001; 22 (2):173-176 - 18.
Kluakamkao G et al. Diagnosis of acute myocardia infarction in sudden unexplained death by a troponin T sensitive rapid assay. Chiang Mai Medical Bulletin. 2004; 43 (2):57-65 - 19.
Davies SJ et al. Investigation of cardiac troponins in postmortem subjects: Comparing antemortem and postmortem levels. The American Journal of Forensic Medicine and Pathology. 2005; 26 (3):213-215 - 20.
Rahimi R et al. Post mortem troponin T analysis in sudden death: Is it useful? The Malaysian Journal of Pathology. 2018; 40 (2):143-148 - 21.
Lai PS et al. Comparison between ante-mortem and post-mortem troponin T. Romanian Journal of Legal Medicine. 2018; 26 :359-362 - 22.
Zhu B et al. Postmortem cardiac troponin T levels in the blood and pericardial fluid. Part 1. Analysis with special regard to traumatic causes of death. Legal Medicine (Tokyo, Japan). 2006a; 8 (2):86-93 - 23.
Zhu B et al. Postmortem cardiac troponin T levels in the blood and pericardial fluid. Part 2: Analysis for application in the diagnosis of sudden cardiac death with regard to pathology. Legal Medicine (Tokyo, Japan). 2006b; 8 (2):94-101 - 24.
Remmer S et al. Cardiac troponin T in forensic autopsy cases. Forensic Science International. 2013; 233 (1-3):154-157 - 25.
Kumar S et al. The effect of elapsed time on cardiac troponin-T (cTnT) degradation and its relation to postmortem interval in cases of electrocution. Journal of Forensic and Legal Medicine. 2015; 34 :45-49 - 26.
Kumar S et al. Temperature-dependent postmortem changes in human cardiac troponin-T (cTnT): An approach in estimation of time since death. Journal of Forensic Sciences. 2016; 61 (Suppl. 1):241 - 27.
Carvajal-Zarrabal O et al. Use of cardiac injury markers in the postmortem diagnosis of sudden cardiac death. Journal of Forensic Sciences. 2017; 62 (5):1332-1335 - 28.
Barberi C, van den Hondel KE. The use of cardiac troponin T (cTnT) in the postmortem diagnosis of acute myocardial infarction and sudden cardiac death: A systematic review. Forensic Science International. 2018; 292 :27-38 - 29.
Cao Z et al. Diagnostic roles of postmortem cTn I and cTn T in cardiac death with special regard to myocardial infarction: A systematic literature review and Meta-analysis. International Journal of Molecular Sciences. 2019; 20 (13):3351 - 30.
Ortmann C et al. A comparative study on the immunohistochemical detection of early myocardial damage. International Journal of Legal Medicine. 2000; 113 (4):215-220. [Accessed: March 7, 2023] - 31.
Martinez Diaz F et al. Biochemical analysis and immunohistochemical determination of cardiac troponin for the postmortem diagnosis of myocardial damage. Histology and Histopathology. 2005; 20 (2):475-481 - 32.
Abdel-Azeem E et al. Medicolegal use of troponin C expression to identify different causes of cardiac deaths at different postmortem intervals. Zagazig Journal of Forensic Medical and Toxicology. 2019; 17 (1):1-9 - 33.
Sabatasso S, Mangin P, Francasso T, Moretti Mx, Docquier M, Djonov V. Early markers of myocardial ischemia and sudden cardiac death. International Journal of Legal Medicine. 2016; 130 :1265-1280 - 34.
Campobasso CP et al. Sudden cardiac death and myocardial ischemia indicators: A comparative study of four immunohistochemical markers. The American Journal of Forensic Medicine and Pathology. 2008; 29 (2):154-161 - 35.
Amin HAA et al. Immuno-histochemistry in the detection of early myocardial infarction (a post mortem study). Egyptian Journal of Forensic Sciences. 2011; 1 :5-12