Prevalence and clinical significance of neutropenia discovered in routine complete blood cell counts: a longitudinal study.

BACKGROUND: Neutropenia, defined as an absolute blood neutrophil count (ANC) <1.5 G L(-1) , may accompany a variety of diseases. However, the clinical significance of neutropenia detected in a routine complete blood cell count is poorly understood. METHODS: Using a primary care resource, comprising more than 370 000 individuals, we assessed the association with a number of previously recognized conditions as well as all-cause mortality in the 4 years following the identification of neutropenia. By matching laboratory data with Danish nationwide health registers, risk estimates were assessed. RESULTS: Neutropenia was observed in approximately 1% of all individuals and was associated dose dependently with viral infections, haematological malignancies (but not autoimmune disorders or solid cancers) and mortality. Neutropenia was particularly associated with HIV, acute leukaemias and myelodysplastic syndromes. Odds ratios [95% confidence interval (CI)] for viral infections were 2.32 (1.84-2.91), 2.80 (2.20-3.57) and 4.77 (3.22-7.07) for subnormal (≥1.5-1.8 G L(-1) ), mild (≥1.0-1.5 G L(-1) ) and moderate-severe (≥0.0-1.0 G L(-1) ) neutropenic individuals, respectively (all P < 0.001). Likewise, odds ratios (95% CI) for haematological malignancies were 3.23 (2.35-4.45), 8.69 (6.58-11.47) and 46.03 (33.98-62.35 ), for the same neutropenia levels, respectively (all P < 0.001). Thus, the lower the ANC, the greater the likelihood of these diseases. The relative risk estimates observed for severe neutropenia corresponded to absolute risks of haematological malignancies and mortality from any cause of 40% and >50%, respectively. CONCLUSIONS: Neutropenia is an ominous sign necessitating careful follow-up. The risk estimates presented here support focusing attention to viral diseases and haematological malignancies when neutropenia is observed.

The Nordic Reference Interval Project 2000: recommended reference intervals for 25 common biochemical properties.

Each of 102 Nordic routine clinical biochemistry laboratories collected blood samples from at least 25 healthy reference individuals evenly distributed for gender and age, and analysed 25 of the most commonly requested serum/plasma components from each reference individual. A reference material (control) consisting of a fresh frozen liquid pool of serum with values traceable to reference methods (used as the project "calibrator" for non-enzymes to correct reference values) was analysed together with other serum pool controls in the same series as the project samples. Analytical data, method data and data describing the reference individuals were submitted to a central database for evaluation and calculation of reference intervals intended for common use in the Nordic countries. In parallel to the main project, measurements of commonly requested haematology properties on EDTA samples were also carried out, mainly by laboratories in Finland and Sweden. Aliquots from reference samples were submitted to storage in a central bio-bank for future establishment of reference intervals for other properties. The 25 components were, in alphabetical order: alanine transaminase, albumin, alkaline phosphatase, amylase, amylase pancreatic, aspartate transaminase, bilirubins, calcium, carbamide, cholesterol, creatine kinase, creatininium, gamma-glutamyltransferase, glucose, HDL-cholesterol, iron, iron binding capacity, lactate dehydrogenase, magnesium, phosphate, potassium, protein, sodium, triglyceride and urate.

Descriptive analytical data and consequences for calculation of common reference intervals in the Nordic Reference Interval Project 2000.

In the Nordic Reference Interval Project (NORIP), data from 102 Nordic clinical chemical laboratories were obtained. Each laboratory reported analytical data on up to 25 of the most commonly used clinical biochemical properties, including results from each of a minimum of 25 reference individuals. A reference material consisting of a liquid frozen pool of serum with values traceable to reference methods (used as the project "calibrator" for non-enzymes to correct reference values) was measured together with other serum pool controls in each laboratory in the same analytical series as the project samples. The data on the controls were used to evaluate the analytical quality of the routine methods. For reference interval calculations, only such reference values on enzymes were accepted that were obtained by applying the International Federation of Clinical Chemistry (IFCC) compatible methods (37 degrees C), while "calibrator"-corrected reference values were used in the cases of non-enzymes. For each property, gender- and age-specific reference intervals were estimated, based on simple non-parametric calculations and using objective criteria to perform partitioning into subgroups. It is concluded that the same reference intervals are applicable in all five Nordic countries. The following descriptive data for the considered properties are presented in the tables: number of measurement values from each country and measurement system, certified/indicative target values for controls, differences between methods and measurement systems together with coefficients of variation, effects of control correction on the measurement values, differences between subgroups as determined by age, gender, country and material, and comparison of the new reference intervals with those presented in standard textbooks. The 25 components involved in this project were (listed in alphabetical order): Alanine transaminase, albumin, alkaline phosphatase, amylase, amylase pancreatic type, aspartate transaminase, bilirubin, calcium, carbamide, cholesterol, creatine kinase, creatininium, gamma-glutamyltransferase, glucose, HDL-cholesterol, iron, iron-binding capacity, lactate dehydrogenase, magnesium, phosphate, potassium, protein, sodium, triglyceride and urate.

Reference individuals, blood collection, treatment of samples and descriptive data from the questionnaire in the Nordic Reference Interval Project 2000.

The rules for recruitment of reference individuals, inclusion and preparation of individuals, blood collection, treatment of samples (and control materials) and analysis at the 102 medical laboratories attending the Nordic Reference Interval Project (NORIP) are given as well as the rules for central exclusion of reference individuals. The individuals (18-91-year-olds) should be evenly distributed on age and gender groups. The 3002 reference individuals who contributed at least one reference value to the finally suggested reference intervals were characterized using the information in the questionnaire. Gender, age and country are the main entries in the tables. Other variables in the cross-tables or figure are height, weight, body mass index, ethnic origin, heredity for diabetes, chronic disease, oestrogens or oral contraceptives, other medication, hard physical activity, previous blood donations, smoking habits, use of alcohol, hours since last meal and time of blood collection (hour, day of week, month, year). The Danes had the highest alcohol consumption and the Icelanders had the highest body mass index. The information in this article may interest potential users of the Nordic Reference Interval Project bio-bank and database (NOBIDA) in which serum, Li-heparin plasma and EDTA buffy coat from the mentioned individuals are stored below -80 degrees C.

Nordic Reference Interval Project Bio-bank and Database (NOBIDA): a source for future estimation and retrospective evaluation of reference intervals.

In the Nordic Reference Interval Project 2000 (NORIP) serum, Li-heparin plasma and EDTA buffy coat were collected at 102 laboratories in 5 Nordic countries from healthy individuals aged 18 years or more and evenly distributed for laboratory, gender and age. Multiple aliquots of these samples from each of about 3000 persons are now stored at the Nordic Reference Interval Project Bio-bank and Database (NOBIDA) at a temperature of below -80 degrees C. The commutable NFKK Reference Serum X with certified values traceable to reference methods and measured in NORIP in the same series as the samples is also available from NOBIDA. Data describing the person and the sample conditions are stored together with analytical results and data describing the measurement systems. The bio-bank along with material and data is administered by the NOBIDA committee on behalf of the NFKK (Scandinavian Society of Clinical Chemistry) to be used by Nordic laboratories for any purpose beneficial to the development of clinical biochemistry in general and particularly for creating reference intervals for other biochemical properties than those established by NORIP. Furthermore, research on the already stored information alone is encouraged. Thus colleagues are now welcome to use this extensive material for research and development in clinical biochemistry.

Estimation of in vivo capillary or venous blood glucose concentration from analysis on stored venous blood or its plasma and use in quality control of near-patient glucose tests.

Glucose concentrations were determined in capillary and venous blood and in venous plasma from 100 consecutive patients referred to an oral glucose tolerance test. The capillary blood was immediately transferred to a haemolysing and glucose stabilizing solution and frozen within 4h. Venous blood was drawn in heparin-sodium fluoride tubes and stored for 0, 4 or 24h at 4 or 20 degrees C. Aliquots of the venous blood were then treated in the same way as the capillary blood. The primary tubes were centrifuged, and aliquots of the plasma were stabilized, as described. All specimens from the same sampling event were analysed in the same analytical series on EBIO compact. Deming linear regression equations y = a + bx were calculated to estimate the glucose concentration in one specimen from that in a differently treated specimen e.g.: B(cB:fPt)---Glucose (0h) = 0.61 + 0.897*P(vB;fPt)-Glucose (24h. 20 C). n = 100, SD(y/x) = 0.25 mmol/L and B(vB) Glucose (0h) = 0.53 + 0.897*P(vB)--Glucose (24h. 20 degrees C), n = 196, SD(y/x) = 0.26 mmol,/L. The non-analytical part of the standard deviation of the ratio between the near-patient test result and its predicted value was 0.038 for both of the above predictions. In conclusion, the analytical (including sampling) variation of near-patient tests of glucose can be assessed by laboratory analysis on mailed, heparin-fluoride stabilized venous blood or on plasma samples made from the mailed blood.

A prospective multipractice study based on antecedent data. Reduced selection and behaviour bias.

A prospective multipractice study based on well-defined data collected by a number of general practitioners (GPs) over a period of time is often used in primary health care research. However, problems of research methodology such as selection bias are common in such studies when GPs either refuse to participate or interrupt their participation. A method is described whereby selection bias is avoided. In the present study, a medical laboratory made a daily record of all abnormal liver function tests requested by GPs over a period of six months. When mailing these results to the GPs, a questionnaire was enclosed in order to obtain a number of clinical data from them. Thirty of 31 possible GPs participated. During the study period, questionnaires concerning 134 patients were mailed, and 118 of these were returned without reminders. Following one reminder the response rate was 100%. Comparing the number of requested tests during the study period with the number requested during a control period before and after the study period (same six month periods of the year), no difference was seen in the pattern of requests. Thus, this method, unbiased with respect to both selection and request-behaviour, is recommended for similar studies.

Epidemiology of abnormal liver function tests in general practice in a defined population in Denmark.

It is common for general practitioners (GPs) to refer patients suspected of impaired liver function for laboratory tests (alkaline phosphatase, lactate dehydrogenase, bilirubin, prothrombin, aspartate aminotransferase). In a prospective multipractice study over a six-month period, including 30 GPs, 55 patients were recorded as having, for the first time, a high level of alkaline phosphatase (AP) as an isolated finding, 14 with an increase of aspartate aminotransferase (ASAT), eight with an increase of both AP and ASAT, three with an increase of ASAT, AP, and bilirubin, two with an isolated increase of lactate dehydrogenase (LDH), one with an increase of ASAT, AP, and bilirubin, combined with a low prothrombin (PP), and, finally, one patient with a low prothrombin in isolation. In most cases the tests were requested because of unspecific symptoms. The most common causes of abnormal test results were neoplasms, alcoholic liver disease, and heart failure. Thirty patients were referred to hospital for further investigations. During the same study period, 50 patients with known abnormal liver function tests were recorded, and the most common causes of these abnormalities were neoplasms, rheumatoid arthritis, and alcoholic liver disease.

Vitamin K to neonates. Peroral versus intramuscular administration.

In a randomized study of 300 infants, the effect of 1 mg of peroral vitamin K given at birth was compared to the same dose given as an intramuscular injection. The combined activity of coagulation factor II + VII + X taken after 48 and before 72 hours after delivery served as the primary endpoint. Prothrombin (antigen) and PIVKA II (acarboxyprothrombin) were also measured. All infants were observed for events of bleeding until discharge from the hospital, normally on the fifth day. No significant differences between the groups in any of the biochemical markers were observed. The 95% confidence limits of the differences were very narrow for all factors. No cases of bleeding were observed. We conclude that administration of 1 mg peroral vitamin K is as efficient as intramuscular administration of the same dose in the prevention of classical hemorrhagic disease of the newborn.

Adjusted-dose intravenous heparin treatment evaluation of an automated and a non automated schedule.

An automated schedule for adjusted continuous intravenous heparin treatment was tested under physician supervision in 19 patients and the results were compared to those obtained in 27 patients treated prior to automation. Both schedules used plasma activated partial thrombo-plastin time (P-APTT) for adjustment of heparin infusion and aimed at the same therapeutic interval for P-APTT (1.5-2 times the value in normal pooled plasma). The quality of treatment was assessed on the basis of clinical information and its ability to keep the P-APTT within or close to the therapeutic interval. The automated schedule reduced the involvement of the physician by 73% without changing the quality of treatment or increasing the consumption of other resources such as the number of P-APTT analyses. Both schedules were able to keep the P-APTT of the patients within the therapeutic interval for about 40% of the treatment time.

Plasminogen activator inhibitors in plasma and platelets from patients with recurrent venous thrombosis and pregnant women.

Using affinity chromatography on lysine Sepharose 4B, a fast-acting tissue plasminogen activator inhibitor (t-PAI) was partially purified from t-PAI-rich plasma from patients with recurrent DVT. Its inhibition of tissue plasminogen activator (t-PA) was demonstrated in functional assays and its reaction with 125I-t-PA was analyzed by autoradiography following SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis). When the t-PAI was mixed with an equimolar concentration of t-PA at 37 degrees C, the half-life of free one-chain and two-chain 125I-t-PA was 1.8 and 0.8 min, respectively. The rate of complex formation between 125I-t-PA and t-PAI was similar both in patient plasma, pregnancy plasma and platelet lysates made from platelet-rich normal, patient and pregnancy plasma. The molecular weights of the complexes between t-PA and the inhibitors in patient plasma and in the different platelet lysates were identical, while that of the inhibitor complex formed in pregnancy plasma was found slightly higher by SDS-PAGE indicating that the pregnancy plasma t-PAI differs from the fast-acting t-PAI found in plasma from thrombotic patients and in platelet lysates.

A functional assay of protein C in human plasma.

A functional assay for protein C in plasma is described in which barium eluates of plasma are incubated with bovine thrombin and rabbit thrombomodulin to activate protein C. The activated protein C solution is added to an activated partial thromboplastin time (APTT) system containing normal plasma and an APTT reagent (Dade ActinR). The prolongation of coagulation time after recalcification in this system is taken as a measure of the anticoagulant activity of protein C. When expressed as per cent of the value in pooled normal plasma, the results obtained by this method in 34 normal controls and in 3 untreated patients with protein C deficiency were very similar to those obtained by radioimmunoassay of protein C. In 2 patients with protein C deficiency and 23 patients without, all on dicoumarol or warfarin treatment, the anticoagulant activity of protein C was less than its antigen concentration. The day to day analytical coefficient of variation (SD/mean) was 12% at the 100% level (n = 12), and 10% at the 25% level (n = 12).

Different types of plasminogen activator inhibitors in plasma and platelets in pregnant women.

The content of tissue plasminogen activator inhibitor (t-PA-I), and of specific pregnancy plasma-PA-I reacting with antibodies to placental-PA-I, was measured in platelet-poor plasma (PPP), platelet-rich plasma (PRP), serum and platelet lysate, both from pregnant women and healthy non-pregnant (male or female) controls. Inhibitory activity was much higher in PPP from pregnant women than in PPP from controls, though it was comparable in platelet lysates both from pregnant women and from controls. The specific pregnancy Plasma-PA-I was present in high concentration in pregnancy PPP, but absent in PPP from the controls and also absent in platelet lysates from both pregnant women and controls. The results show that the specific pregnancy plasma-PA-I is not identical to either the PA-I in platelet lysate or to the PA-I in PPP from non-pregnant controls.

Rates of synthesis of prealbumin and retinol-binding protein during acute inflammation in the rat.

The rates of synthesis of prealbumin (PA), retinol-binding protein (RBP), and other plasma proteins were measured in primary monolayer cultures of rat hepatocytes isolated from normal rats and from rats 18 h after induction of an inflammatory reaction by subcutaneous injection of croton oil. The inflammatory pattern of protein synthesis seemed to persist in the isolated hepatocytes for 1-2 days. This pattern included significantly decreased rates of synthesis of PA. The rate of synthesis of RBP was probably also decreased, but significantly less than the rate of PA synthesis. The results support the idea that it is mainly the decreased rate of PA synthesis which is responsible for the decreased plasma concentration of PA, and its ligand RBP and retinol during inflammation.

Prealbumin in Swedish patients with senile systemic amyloidosis and familial amyloidotic polyneuropathy.

A prealbumin (PA)-like protein was demonstrated in amyloid fibrils both from a patient with senile systemic amyloidosis (SSA) and from a patient in northern Sweden with familial amyloidotic polyneuropathy (FAP). The investigated properties of this protein were similar in the two types of fibrils. The protein had molecular weight, antigenic determinants, and at least one cysteinyl residue in common with the subunit of normal PA. In contrast to normal PA, it contained disulphide-linked subunits and was to some extent bound to the fibril via the cysteinyl residues. The noncovalent forces between its subunits were weaker than in normal PA. It constituted part of the previously described AScl protein of the SSA fibrils. Proteins with lower molecular weight than the PA monomer were major proteins in both SSA and FAP fibrils. These proteins had similar properties in the two kinds of fibril and may be derived from PA. PA in serum from Swedish patients with FAP and SSA was normal with regard to the isoelectric pH of the monomers and tetramers.

The increased negative charge of prealbumin in cerebrospinal fluid is acquired in vitro by oxidation of the cysteinyl residues without formation of disulphides.

The reported charge difference between prealbumin (PA) in cerebrospinal fluid (CSF) and PA in plasma has been traced to the cysteinyl residues. In freshly drawn CSF and plasma, PA had the same electrophoretic mobility at pH 8.6 and the same microheterogeneity by electrofocusing under denaturing and nondenaturing conditions. No differences in molecular weight were detected on sodium dodecyl sulphate polyacrylamide slab gel electrophoresis between PA isolated from stored CSF and PA isolated from serum. In fresh samples of CSF and plasma almost all half-cystine residues of PA reacted with 5,5'-dithio bis (2-nitrobenzoic acid) (DTNB). During storage of CSF and plasma the single cys-10 in the PA subunits acquired an extra negative charge and became unreactive towards DTNB due to oxidation mainly without formation of disulphides. This reaction proceeded faster in CSF than in plasma and explains the reported lower isoelectric pH and the higher electrophoretic mobility of PA in CSF compared to the corresponding plasma.

Factors affecting the concentration of free holo retinol-binding protein in human plasma.

The total concentrations of retinol, retinol-binding protein and prealbumin were determined in plasma from twenty healthy men and sixty patients with various inflammatory conditions. These concentrations were all strongly correlated to each other and lower in the patient group. The concentration of free (not prealbumin-bound) holo retinol-binding protein, the presumed 'active' supplier of retinol to the tissues, was calculated. It was found not to be decreased in the patient group. Of the measured total concentrations and their possible ratios in the whole material, the retinol/prealbumin ratio showed the strongest correlation to the concentration of free holo retinol-binding protein. The importance of the concentration of free holo retinol-binding protein for the vitamin A supply to the cells was supported by calculations on data from the literature showing that this concentration better than the above-mentioned total concentrations distinguished between patients with normal and abnormal dark adaptation ability.