Eva Landberg

To mix or not to mix venous blood samples collected in vacuum tubes

Abstract Background: There are recommendations to mix venous blood samples by inverting the tubes immediately after venipuncture. Though mixing allows efficient anticoagulation in plasma tubes and fast initiation of coagulation in serum tubes, the effect on laboratory analyses and risk of haemolysis has not been thoroughly evaluated. Methods: Venous blood samples were collected by venipuncture in vacuum tubes from 50 patients (10 or 20 patients in each group). Four types of tubes and 18 parameters used in routine clinical chemistry were evaluated. For each patient and tube, three types of mixing strategies were used: instant mixing, no mixing and 5 min of rest followed by mixing. Results: Most analyses did not differ significantly in samples admitted to different mixing strategies. Plasma lactate dehydrogenase and haemolysis index showed a small but significant increase in samples omitted to instant mixing compared to samples without mixing. However, in one out of twenty non-mixed samples, activated partial thromboplastin time was seriously affected. Conclusions: These results indicate that mixing blood samples after venipuncture is not mandatory for all types of tubes. Instant mixing may introduce interference for those analyses susceptible to haemolysis. However, tubes with liquid-based citrate buffer for coagulation testing should be mixed to avoid clotting.

Disialo–trisialo bridging of transferrin is due to increased branching and fucosylation of the carbohydrate moiety

BACKGROUND Carbohydrate deficient transferrin (CDT) is used for detection of alcohol abuse and follow-up. High performance liquid chromatography (HPLC) of transferrin glycoforms is highly specific for identification of alcohol abuse, but unresolved disialo- and trisialotransferrin glycoforms sometimes makes interpretation difficult. The cause of this phenomenon is unknown, cannot be explained by genetic variants of transferrin, but seems to be associated with liver disease. METHODS Nineteen serum samples showing di-tri bridging when analyzed by HPLC were collected. Transferrin was purified by affinity chromatography, and N-linked oligosaccharides were released enzymatically. The N-glycans were further analyzed by high performance anion-exchange chromatography with pulsed amperometric detection and MALDI-TOF mass spectrometry. RESULTS The HPLC-analysis showed three different types of glycoform patterns. The N-glycans from fifteen samples showed patterns with increased number of triantennary structures containing one or two fucose residues. One sample contained an increased amount of triantennary glycans without fucose. Three samples showed a glycosylation pattern similar to normal transferrin. CONCLUSIONS The di-tri bridging phenomenon was associated with alterations in transferrin glycosylation in the majority of cases. Transferrin contained a higher extent of triantennary and often fucosylated N-linked oligosaccharides. These results may be important in future diagnostic approaches to liver diseases.

Temperature effects in high-performance anion-exchange chromatography of oligosaccharides.

High-performance anion-exchange chromatography (HPAEC) with pulsed amperometric detection has been widely used for analysis of mono-, oligo- and polysaccharides. Many factors that affect separation of carbohydrates by HPAEC have been evaluated, however effect of temperature has not been carefully studied. In the present study, neutral and sialylated oligosaccharides from human milk and different types of N-linked oligosaccharides were analysed by HPAEC at temperatures ranging from 13 to 30 degrees C. N-Acetyl neuraminic acid, Galacturonic acid and stachyose were also analysed since they have been used as internal standards when analysing various oligosaccharides by HPAEC. All oligosaccharides showed decreased retention times with increased temperature. Even small differences (i.e. +/- 5 degrees) resulted in considerable changes in retention times. In addition, individual oligosaccharides showed relative changes in retention time with increased temperature. By changing the temperature, a switch in elution of order of individual oligosaccharides were sometimes found. These results show that retention times relative to an internal standard cannot be used for oligosaccharide identification unless temperature is carefully controlled. Regulation of temperature is also a valuable tool in achieving optimal separation of oligosaccharides by HPAEC.

Point-of-Care Troponin T Testing in the Management of Patients with Chest Pain in the Swedish Primary Care

Objective. To investigate the diagnostic accuracy and clinical benefit of point-of-care Troponin T testing (POCT-TnT) in the management of patients with chest pain. Design. Observational, prospective, cross-sectional study with followup. Setting. Three primary health care (PHC) centres using POCT-TnT and four PHC centres not using POCT-TnT in the southeast of Sweden. Patients. All patients ≥35 years old, contacting one of the primary health care centres for chest pain, dyspnoea on exertion, unexplained weakness, and/or fatigue with no other probable cause than cardiac, were included. Symptoms should have commenced or worsened during the last seven days. Main Outcome Measures. Emergency referrals, patients with acute myocardial infarctions (AMI), or unstable angina (UA) within 30 days of study enrolment. Results. 25% of the patients from PHC centres with POCT-TnT and 43% from PHC centres without POCT-TnT were emergently referred by the GP (P = 0.011 ). Seven patients (5.5%) from PHC centres with POCT-TnT and six (8.8%) from PHC centres without POCT-TnT were diagnosed as AMI or UA (P = 0.369). Two patients with AMI or UA from PHC centres with POCT-TnT were judged as missed cases in primary health care. Conclusion. The use of POCT-TnT may reduce emergency referrals but probably at the cost of an increased risk to miss patients with AMI or UA.

Serum prolactin and macroprolactin in heart failure: no relation to established laboratory or clinical parameters

Background A few smaller studies have reported that the prolactin concentration is elevated in connection with heart failure. As heart failure is combined with disturbances of several biological systems any or all of which may also influence prolactin concentrations, we wanted to evaluate the relation of prolactin to prognosis in elderly patients. Methods A total of 462 elderly patients from a primary health-care centre, all with symptoms of heart failure, were included. In addition to clinical examination including echocardiography, concentrations of prolactin, macroprolactin, C-reactive protein, thyroid-stimulating hormone and N-terminal pro B-type natriuretric peptide (NT-proBNP) were measured. Patients were then followed for 10 y, and all incidents of cardiovascular mortality were registered. Results After excluding patients with macroprolactin, hyperprolactinaemia was found in 3.7% of the patients. There were no differences in prolactin concentrations or in the frequency of macroprolactin between patients with heart failure and those with normal cardiac function, defined as left ventricular ejection fraction of at least 50%. No significant correlation could be found between NT-proBNP and prolactin. Neither could any association be found between cardiovascular mortality and prolactin concentration during 10 y of follow-up. Conclusions Prolactin concentrations were not associated with cardiovascular mortality or any clinical or biochemical marker of heart failure. Macroprolactin was found in similar frequency among patients with and without heart failure, and showed no correlation with mortality risk.

Effects of prolactin on platelet activation and blood clotting

Abstract Increased levels of prolactin often coincide with an increased risk for thromboembolic events, but it is unclear whether a direct causal relation exists. Our aim was to examine the effect of prolactin on platelet function. In addition to using recombinant prolactin for experiments in vitro, we analyzed platelet function by flow cytometry in a group of 13 females with hyperprolactinaemia and 18 healthy female controls. Platelet activation was measured by P-selectin expression and by the amount of platelet-bound fibrinogen after stimulation with adenosine di phosphate (ADP), collagen-related peptide and the protease activated receptor (thrombin receptor) (PAR)-activating peptides PAR4-AP and PAR1-AP. Free oscillation rheometry was used to measure clotting time in whole blood. No significant effect on platelet activation or clotting time could be seen in in vitro experiments by adding recombinant prolactin. However, significantly lower P-selectin expression was found in the hyperprolactinemic group when platelets were activated by ADP (5 and 10 μM) or PAR4-AP. The expression of fibrinogen did not differ between the two groups for any of the activators used. For all samples, inverse significant correlations between P-selectin expression and prolactin concentration were found for both 5 μM ADP (r = − 0.61, p < 0.01), 10 μM ADP (r = − 0.62, p < 0.001) and PAR4-AP (r = − 0.69, p < 0.001). Thrombin cleavage of recombinant prolactin resulting in a 16 kDa C-terminal fragment did not alter the P-selectin expression upon activation. We found an indirect inhibitory effect of prolactin on platelets in hyperprolactinemic patients, suggesting that prolactin might have a protective role in thromboembolic disease.