Sujiva Ratnaike

The accuracy of cystatin C and commonly used creatinine-based methods for detecting moderate and mild chronic kidney disease in diabetes

Background  The accuracy of measuring serum cystatin C levels for detecting various stages of chronic kidney disease (CKD) in diabetes is still unclear.

Capillary isoelectric focusing of haemoglobin variants in the clinical laboratory

For capillary isoelectric focusing (CIEF) to be accepted in the clinical laboratory, it must be reproducible and cost effective. The advent of polyAAEE coated capillaries (Bio-Rad Laboratories, Hercules, CA, USA) has provided the means of obtaining over 100 runs per capillary, something which previously had not always been possible with coated capillaries. Using the Clinical Data Management computer program on the BioFocus 2000 Capillary Electrophoresis System (Bio-Rad Laboratories), we have used a one-step salt mobilization to achieve focusing of haemoglobin variants. Washed red cells are diluted, haemolyzed and separated in the capillary at 8 kV using 1.3% Pharmalyte ampholytes (pH 6.6-7.7/pH 6-8 2:1) in 0.40% methylcellulose. The separated haemoglobins were detected by adsorption at 280 nm. Using published values of haemoglobin variants, we investigated the use of pI markers to confirm the pI of haemoglobin variants detected. CIEF, though more expensive than capillary electrophoretic separations of haemoglobin variants, has greater resolution due to the fact that the separation of variants from pI 6.95 to 7.42 occurs over 4 min, whereas the electrophoretic separation is over 60 s. CIEF is quicker than gel IEF, and shows real-time results as the sample is being processed. The potential for CIEF in the clinical laboratory is not limited to haemoglobin variants, and the technique should become increasingly popular in the near future.

Cystatin C for estimation of glomerular filtration rate in patients with spinal cord injury

Background: Serum creatinine is not a satisfactory marker of glomerular filtration rate (GFR) in patients with spinal cord injury (SCI) who have varying degrees of muscle atrophy. In contrast to serum creatinine, serum cystatin C, a 13-kDa protein, is not affected by muscle mass and is therefore potentially a useful marker of GFR in patients with SCI. In addition, cystatin C is not dependent on sex or age and is not secreted by the renal tubule. Aim: We assessed serum cystatin C as a surrogate marker of GFR in SCI patients. Methods: Cystatin C was analysed using a particle-enhanced immunonephelometric assay (Dade Behring) in serum samples sent for routine measurement of creatinine (64 patients) and creatinine clearance (27 patients) from patients in the Spinal Unit of the Austin Health. We compared these results with serum cystatin C of 57 non-SCI patients who had had a creatinine clearance measurement during the study period. Results: In patients with SCI, the reciprocal of cystatin C had a stronger correlation (r = 0·48, P<0·01) with creatinine clearance than the reciprocal of serum creatinine (r = 0·25, P<0·19). Further, the value of serum creatinine was much lower for a given creatinine clearance in SCI patients than in non-SCI patients; the serum cystatin C concentrations were equivalent. Conclusion: The serum cystatin C is a convenient and more reliable surrogate marker of GFR than serum creatinine and will enable early detection of renal impairment. We need to confirm this finding with a larger study, including comparison with an accepted gold standard for GFR.

Capillary Electrophoresis of Hemoglobin

Abstract Capillary electrophoresis (CE) has been used in a variety of in-house capillary isoelectric focusing (CIEF) and capillary zone electrophoresis (CZE) assays for the detection of hemoglobin (Hb) variants and the quantitation of HbA2 and HbF. A commercial kit has also been produced for the analysis of hemoglobin variants and thalassemia screening. Though CE methods have been shown to be able to detect many variants, final identification of the variant needs specialized testing such as DNA technology. Over the past 2 years, many instruments that had been used for these hemoglobin variant screening and thalassemia assays have been withdrawn from sale. Although CE HbA1c analysis is available, it cannot compete in turnaround time or cost with automated HPLC commercial instruments that give accurate HbA1c results in 3 or 4 minutes. Hence we do not anticipate a bright future for the analysis of hemoglobin by CE.

N -Acetylcysteine does not artifactually lower plasma creatinine concentration

BACKGROUND All randomized controlled trials of N-acetylcysteine (NAC) in contrast media-induced nephropathy used creatinine as a marker of renal function. However, it has been suggested that NAC may lower plasma creatinine levels independent of any effects on glomerular filtration rate (GFR). METHODS At a tertiary hospital 110 cardiac surgical patients were randomly allocated to peri-operative infusion of NAC (300 mg/kg over 24 h, N = 30) or placebo (N = 80). We compared the plasma concentrations of creatinine, cystatin C and urea, the plasma creatinine/plasma cystatin C ratio and the estimated GFR at baseline and at 24 and 72 h after commencement of the infusion. We measured urinary creatinine concentration at 24 h. RESULTS At baseline, the plasma creatinine/plasma cystatin C ratio did not differ between the NAC and placebo group (0.90 versus 0.92; P = 0.94). There was no significant difference in the plasma creatinine/plasma cystatin C ratio for the NAC and placebo group either during or after NAC infusion at 24 h (1.03 versus 1.00; P = 0.78) and 72 h (0.94 versus 0.89; P = 0.09). Those allocated to NAC showed no difference in urinary creatinine excretion when compared to placebo (P = 0.24). CONCLUSIONS The results of our study do not demonstrate that NAC artifactually lowers creatinine measured using the Jaffé method. (ClinicalTrials.gov, NCT00332631, NCT00334191).

The diagnosis and follow-up of porphyria

&NA; This review details an approach to the biochemical diagnosis and follow‐up of porphyria. We discuss the problems of diagnosis of both symptomatic patients suspected of porphyria and patients being investigated because of a family history of porphyria. High performance liquid chromatography plays a major role in the investigation of these patients. Molecular biology is emerging as a useful tool in further defining this group of diseases.

Single and serial measurements of cardiac troponin I in asymptomatic patients on chronic hemodialysis.

AIMS Coronary artery disease is the major cause of death in patients with end-stage renal failure on dialysis. This study aimed to assess the predictive value of a single cardiac troponin I (cTnI), and also the kinetics of serial values. METHODS Since cTnI is a potential biomarker of cardiac outcome, the present study examined single cTnI measurements (n = 88 patients) and its predictive value for future cardiac events, and a kinetic substudy of serial weekly cTnI measured for 8 weeks (n = 57) in a group of patients on hemodialysis. RESULTS Single cTnI measurements: 9 patients (10.2%) had a detectable cTnI at baseline and 79 patients (89.8%) had a negative baseline cTnI. There were no significant differences in age, sex, history of ischemic heart disease, diabetes, smoking or dyslipidemia between patients with detectable and negative cTnI. At the end of 9 months, the rate of combined primary endpoints, which included myocardial infarction, cardiac death and cardiac revascularization, was significantly higher in the patients with a detectable baseline cTnI (55.6%), compared to patients with a negative cTnI (6.3%) (p = 0.0007). Serial weekly cTnI measurements: significant fluctuations in cTnI were noted over time; 27% of patients with an undetectable cTnI measured at baseline had subsequent detectable levels in the serial follow-up. CONCLUSION A single detectable cTnI in asymptomatic patients on hemodialysis defines patients at high risk of future cardiac events. However, the incidence of detectable cTnI levels is markedly increased when serial weekly measurements are performed. The clinical significance of detectable serial measurements of cTnI is the focus of ongoing studies.

B-type Natriuretic Peptides Strongly Predict Mortality in Patients Who Are Treated with Long-Term Dialysis

BACKGROUND AND OBJECTIVES Left ventricular abnormalities contribute to cardiovascular disease in patients with chronic kidney disease and may be detected by measurement of B-type natriuretic peptide in serum. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS In a prospective cohort study of predialysis patients, patients who were on dialysis, and kidney transplant recipients, serum was collected and assayed for both B-type natriuretic peptide and its N-terminal fragment. Median levels were compared using nonparametric tests, and predictors of B-type natriuretic peptide were determined by linear regression. Survival analysis and Cox regression were performed to examine the association of levels of B-type natriuretic peptide with cardiovascular events and death. RESULTS Levels of B-type natriuretic peptide were highest in patients who were on dialysis. Patients who were receiving dialysis and had known cardiovascular disease, were not on the waiting list for kidney transplantation, or had left ventricular systolic dysfunction on echocardiography had significantly higher levels of B-type natriuretic peptide than patients without these characteristics. Glomerular filtration rate was an important predictor of B-type natriuretic peptide levels for patients who were not on dialysis (predialysis and renal transplant recipients). Left ventricular systolic dysfunction predicted B-type natriuretic peptide levels in patients who were on dialysis. Both forms of B-type natriuretic peptide were associated with a two- to three-fold increased risk for death in patients who were on dialysis. CONCLUSIONS Levels of B-type natriuretic peptide are greatest in patients who are on dialysis and have cardiovascular comorbidities and are strong predictors of death.

ALPHA1-Antitrypsin phenotypes in homosexual men

&NA; The alpha1‐antitrypsin (AAT) phenotype was determined by isoelectric focusing in 215 male homosexuals and compared with those in 208 male heterosexuals. The incidence of abnormal phenotypes was 16.3% in the homosexual group which was significantly different (p<0.03) than the 8.7% in the heterosexual group. There was no difference in the phenotype distribution between homosexuals who were anti‐human immunodeficiency virus reactive and those who were non‐reactive. It suggests that investigation into the interplay of factors associated with homosexuality could include genetic as well as psychological and social factors.

Serial increased cardiac troponin T predicts mortality in asymptomatic patients treated with chronic haemodialysis

Background A single detectable cardiac troponin predicts mortality in patients treated with dialysis. There are limited data on changes in troponin concentration over time and the clinical implications of serial troponin measurement. Methods Serial cardiac troponin T (cTnT) was assayed five times over 12 months in a prospective cohort study of patients with end-stage kidney disease treated with haemodialysis. A concentration of cTnT ≥0.04 μg/L was considered increased. Mortality and cardiovascular events were analysed by survival analysis, according to the serial troponin results. Results From 100 patients who provided a baseline sample for cTnT, 81 completed five serial measurements. The analysis of patients who completed serial cTnT measurements demonstrated that 28 patients (35%) had normal cTnT concentrations in all five samples, 20 patients (24%) had between one and four increased cTnT measurements and 33 patients (41%) had increased concentrations of cTnT in all five samples. The 1.7-y patient survival was 100%, 90% and 78% for patients with zero, one to four, or five out of five concentrations of cTnT increased, respectively (P = 0.037), and the corresponding cardiovascular event-free survival was 100%, 91% and 78%, respectively (P = 0.027). Conclusions Serial measurements of cTnT concentration were frequently increased in patients receiving haemodialysis. The number of abnormal measurements over time predicted mortality and cardiovascular adverse events.