Laboratory control of oral anticoagulant therapy: preservation of prothrombin time specimens using a polypropylene collection system

Previous studies have shown a marked time and temperature dependent shortening of the prothrombin time (PT) when blood is exposed to borosilicate (glass) or siliconized borosilicate tubes. Current recommendations are that samples for PT estimation should be tested within 2 h of collection. In this study using polypropylene collection tubes, blood obtained from 30 patients on oral anticoagulant therapy showed no significant change in International Normalized Ratio (INR) value after 24 h storage--either at 4 degrees C or room temperature. After 48 h. changes in INR values from refrigerated samples were still clinically insignificant. After 48 h storage at room temperature, however, a minority of samples showed an increase in INR value which may be of clinical importance. The range of INRs studied was 1.0-9.1. In a second evaluation, replicate specimens from 22 orally anticoagulated patients with INRs ranging from 1.0 to 9.6 showed no significant change after 24 h at either temperature--even when samples had been subjected to 30 min of gentle agitation prior to storage and analysis. Overall, the results indicate that when polypropylene collection tubes are used, prothrombin time specimens can be successfully preserved for up to 24 h at room temperature or up to 48 h when refrigerated.     

Recovery of nosocomial fecal flora from frozen stool specimens and rectal swabs: comparison of preservatives for epidemiological studies

The recovery of antibiotic-susceptible and -resistant aerobic Gram-negative bacilli from stool specimens and from mock rectal swabs after freezing (-20 degrees C) for as long as 4 weeks was studied using three preservatives: Cary-Blair (CB) transport medium, buffered glycerol saline (BGS), and Para Pak C&S solution (CS). In addition, the recovery of enterococci from rectal swabs was investigated after storage of swabs in Stuart's transport media at 4 degrees C as long as 4 weeks. The log10 decreases in bacterial counts from seeded stool suspensions frozen in BGS were 0.64 (i.e., fourfold) and 1.16 after 1 and 4 weeks, respectively, which were significantly less (p < .05) than 1 and 4 week decreases following freezing in CB (1.57 and 2.85) or in CS (1.50 and 2.45). The recovery of Gram-negative bacilli from patients' rectal swabs preserved in BGS was consistent with the results of the experiments with seeded stool suspensions. There was no detectable decrease in recovery of enterococci from rectal swabs stored at 4 degrees C. BGS performed well as a preservative for freezing stool specimens or rectal swabs for later recovery of nosocomial Gram-negative bacilli; enterococci survived well in refrigerated rectal swab specimens.     

Effect of different storage temperatures, sample collection procedures and immunoassay methods on osteocalcin measurement

The apparent instability of measured osteocalcin has been reported as method-dependent and related to preanalytical variables such as storage temperature, and the use of anticoagulants and protease inhibitors. The aim of this study was to determine a sample collection procedure which minimised osteocalcin degradation. Blood samples from five normal individuals were collected with or without anticoagulants and protease inhibitors (heparin, EDTA, or heparin and aprotinin) and stored at 4 degrees C, -20 degrees C or -70 degrees C for up to 7 days, 28 days and 90 days respectively. Osteocalcin was measured by both a monoclonal EIA specific for intact osteocalcin and a bovine polyclonal RIA. Osteocalcin concentrations in serum and EDTA-treated samples significantly decreased by 40% (P < 0.001) with the ELISA and 72% (P < 0.001) with the RIA after 7 days storage at 4 degrees C. Similar falls were documented in these samples when stored at -20 degrees C and -70 degrees C and measured by the ELISA. Minimal changes in osteocalcin immunoreactivity were observed in either assay when heparin-treated plasma with or without aprotinin was stored at -20 degrees C or -70 degrees C for up to 90 days. The apparent instability of measured osteocalcin can be minimised using these conditions.     

Meat toughening does not occur when rigor shortening is prevented

The objective of this experiment was to test the hypothesis that meat toughening during the first 24 h postmortem results from sarcomere shortening during rigor mortis development. Eleven market-weight lambs were used to measure changes in shear force of clamped longissimus during rigor development. Within 15 min of exsanguination, while attached at both ends, each longissimus was separated from the vertebrae body and clamped between three sets of metal plates to prevent muscle shortening (six clamped sections per lamb). Five of the clamped sections were placed at -1.1 degrees C for 0, 3, 6, 12, or 24 h. After storage at their respective times at -1.1 degrees C, the samples were placed at -30 degrees C for 90 min and then at -5 degrees C for 8 d. The sixth section (168-h section) was stored at -1.1 degrees C for the first 24 h, at 4 degrees C for 144 h, and then treated the same as other sampling times. Sections were sampled for pH, sarcomere length, shear force, and Western blot analyses before and after storage at -5 degrees C. Shear force values were the same (P > .05) from 0 to 24 h (4.5 kg at 0 h to 4.9 kg at 24 h) then declined (P < .05) to 3.3 kg at 168 h postmortem. As evident by lack of statistical difference in the sarcomere lengths, we were successful in holding the muscle length constant. Western blot analyses of nebulin, vinculin, and troponin-T indicated that minimum degradation occurred through 12 h, was slightly increased by 24 h, and was relatively extensive by 168 h postmortem. Although limited proteolysis occurred during storage at -5 degrees C for 8 d, this by itself had no effect on shear force. Results indicate that shear force values do not increase during rigor development when muscle is prevented from shortening; thus, the toughening that occurs during the first 24 h of slaughter is most likely due to sarcomere shortening.     

A novel isoform of the neurofibromatosis type-1 mRNA and a switch of isoforms during murine cell differentiation and proliferation

There are different recommendations for the handling of blood samples for analyses of the kallikrein-kinin or complement system, respectively. C1 inhibitor (C1-INH) takes a crucial part in both systems. In order to establish recommendations for blood specimen collection and transport for making the diagnosis of hereditary angioedema (HAE), the effect of time, temperature and different additives on C1-INH function and antigen was determined. We used blood samples from normals and patients suffering from HAE type I. Plasma containing EDTA, heparin, sodium citrate or polybrene-EDTA, and serum were assayed after incubations at 4 degrees C or 37 degrees C for 6 or 24 h. In addition, pooled serum was incubated for up to 5 days at room temperature. A modest decrease in C1-INH function was observed as an effect of storage-time in samples from normals (p = 0.039) and a substantial decrease was seen for the HAE patients (p = 0.0002). No significant effect of temperature (4 degrees C or 37 degrees C) was found. Clotting did not reduce C1-INH activity. Plasma containing heparin or polybrene interfered with the functional assay, yielding falsely high and low values, respectively. C1-INH functional assay performed within 24 h in serum, EDTA-treated or citrated plasma discriminated well between HAE patients and normals. This was also the case for serum kept at room temperature for up to 5 days, although a modest fall in C1-INH function was seen in the incubation period. For practical purposes we recommend serum as the sample of choice, preferably received within 48 h.     

Effects of specimen collection, processing, and storage conditions on stability of human immunodeficiency virus type 1 RNA levels in plasma

To define the optimal blood collection parameters for plasma human immunodeficiency virus type 1 (HIV-1) viral load testing, plasma HIV-1 RNA levels were quantitated with the NASBA HIV-1 RNA QT System from blood specimens that were collected, processed, and stored under a variety of conditions that might have affected HIV-1 RNA stability. We determined that when whole blood was processed within 2 h of specimen collection the levels of HIV-1 RNA detected in EDTA-, heparin-, and acid citrate dextrose (ACD)-anticoagulated plasma samples were comparable. The levels of HIV-1 RNA in serum specimens (mean = 4.126 log units) were significantly lower (P < 0.01) than the levels in corresponding plasma samples (mean = 4.501 log units). One cycle of freeze-thaw (-70 degrees C) did not significantly reduce the level of HIV-1 RNA detected in EDTA-, heparin-, or ACD-anticoagulated plasmas. The EDTA-anticoagulated plasmas showed the smallest decrease in HIV-1 RNA copies (0.050 log units). HIV-1 RNA levels decreased over a 6-month time period in serum as well as in EDTA-, ACD-, and heparin-anticoagulated plasmas stored at -70 degrees C. However, the only significant decreases were for serum (mean decrease = 0.317 log units) and heparin-anticoagulated samples (mean decrease = 0.384 log units). A comparison of the levels of HIV-1 RNA in cell-free plasma collected in VACUTAINER EDTA Plasma Preparation Tubes and in standard VACUTAINER EDTA tubes determined that HIV-1 RNA levels were stable for up to 30 h after collection when stored at either room temperature (mean standard deviation [SD] = +/- 0.101 log units) or at 4 degrees C (mean SD = +/- 0.102 log units) as cell-free plasma or as EDTA-anticoagulated whole blood (mean SD = +/- 0.109 log units). These data indicate that EDTA-anticoagulated plasma is the most suitable and stable matrix for HIV-1 RNA quantitation.     

Reproducibility of electronic cell counts in milk; a study of 5 further factors

This paper is a sequel to a previous one in which a number of factors likely to influence the accuracy of counting somatic cells in milk was assessed; in the present work the effects of 5 other factors are investigated. In a study of storage time and temperature of milk samples fixed in formalin, a significant increase in cell count occurred after 5-7 d when samples were stored at room temperature (17-23 degrees C), compared with those maintained at 4 degrees C. When manual and mechanical mixing of fixed samples were compared only marginal differences in cell counts were observed. An increase in cell counts followed manual dilution of milk samples in comparison with automatic dilution. The temperature of samples prepared for counting was also studied and no significant variations occurred between mean temperatures of 12-7 and 32-9 degrees C. The final factor evaluated was that of holding time before counting; using 4 cell-count levels it was observed that counts were acceptable up to 1 1/2 h.     

Comparison of manual and automated cell counts in EDTA preserved synovial fluids. Storage has little influence on the results

OBJECTIVE: To determine the precision and agreement of synovial fluid (SF) cell counts done manually and with automated counters, and to determine the degree of variability of the counts in SF samples, kept in the tubes used for routine white blood cell (WBC) counts--which use liquid EDTA as anticoagulant--at 24 and 48 hours at 4 degrees C, and at room temperature. METHODS: To determine precision, cell counts were repeated 10 times--both manually and by an automated counter--in a SF sample of low, medium, and high cellularity. The variances were calculated to determine the interobserver variation in two manual (M1,M2) and two automated cell counts (C1,C2). The agreement between a manual (M1) and automated counter (C1) results, was analysed by the Bland and Altman method and the difference against the mean of the two methods was plotted. Then, the mean difference between the two methods was estimated and the standard deviation of the difference. To determine the effects of storage, SF samples were kept in a refrigerator at 4 degrees C, and at room temperature; cell counts were done manually (M1) and automatically (C1) at 24 and 48 hours and the changes analysed by the Bland and Altman method. The variances were compared using an F test. RESULTS: (1) Precision. With the manual technique, the coefficients of variation were 27.9%, 14%, and 10.7% when used for counting the SF with low (270), medium (6200), and high cellularities (25,000). With the automated technique the coefficients of variation were 20%, 3.4%, and 2.9% in the same SF samples. In the fluids of medium and high cellularity, the variances of the automated cell counts were significatively lower (F test, p < 0.002) than those of the manual counts. (2) Interobserver variation. The variance between C1 and C2 (25 SF) was significatively lower (F test, p < 0.002) than that of the manual counts (41 SF). (3) Agreement between the two techniques (100 SF). For cellularities above 2000 cells/mm3, the manual method gave results between +10% to -34% of the results obtained by the coulter. For cellularities below 2000 cells/mm3, manual cell counts were between +60 to -1280 cells/mm3 of those obtained by the automated counter. (4) Influence of storage. The coulter counts of SF samples preserved at 4 degrees C showed less variance (F test, p < 0.05) than the manual counts. The worst results were obtained in manual counts of SF samples kept at room temperature; these samples at 48 hours showed a variation between -47% to 42% of the initial results. CONCLUSIONS: Automated cell count of the SF offers advantages: it gives higher precision and consumes less time. The stability of the samples preserved in the EDTA tubes used for routine WBC counts is of additional interest, because if delay cannot be avoided, the results of the WBC counts are still accurate at 24 and even at 48 hours, at least for clinical purposes.     

Flow-cytometric analysis of reticulocytes in normal cord blood

We performed precise reticulocyte counts and a reticulocyte maturation study according to ribosomal RNA (rRNA) content, in normal cord blood. For this purpose we analyzed 35 cord blood specimens corresponding to a mean gestational age of 39.2 weeks (range 38-41). In all specimens complete blood counts were performed with the H*1 (Technicon, Bayer) analyzer. Reticulocyte maturation study and counts were conducted by the R-1000 (Sysmex) reticulocyte analyzer. We obtained the following measurements (mean +/- 1 SD): reticulocyte percentage 3.11 +/- 0.75% and reticulocyte absolute count 137.3 +/- 33.3 x 10(9)/l. There was no correlation between the reticulocyte counts and the duration of gestation or the type of labor (normal, forceps assisted or cesarean section). Reticulocyte subpopulations with different rRNA content and maturation were expressed as reticulocytes with high (HFR), median (MFR) and low (LFR) fluorescence of the fluorochrome auramine bound to rRNA. Normal cord values were: HFR% = 13.6 +/- 2.4, MFR% = 22.5 +/- 2.4 and LFR% = 63.9 +/- 4.3. Reference maturation subpopulations were assessed for comparison in 180 samples of healthy adults (90 males and 90 females) and were found HFR% = 1.0 +/- 0.8, MFR% = 9.7 +/- 3.3 and LFR% = 89.2 +/- 3.4. The HFR% and MFR% were significantly higher while LFR% was significantly lower (p < 0.001) in cord compared to adult blood, denoting a shift to more immature reticulocyte forms. The above values are provided as normal reference data of the reticulocyte maturation profile in full-term cord blood.     

The protection of creatine kinase MM sub-bands by EDTA during storage

We examined the effect of a 5 mmol/l concentration of EDTA on the stabilization of the five serum creatine kinase MM isoenzymes, resolved by thin-layer isoelectric focusing. In patient sera, total CK and CK-MB activities were stable during storage of the samples for two months at 4 degrees C even in the absence of EDTA. However, EDTA stabilized the labile MM and MM1 sub-bands, which are the first to appear in the blood after the release from the damaged tissue and its addition to blood samples intended for determining the MM sub-band pattern is recommended. The stabilizing effect of EDTA was emphasized at higher temperatures. EDTA protected the CK-MM pattern in myocardium extracts made in normal serum and incubated at 37 degrees C during 40 h, but was unnecessary when myocardium was homogenized in heat-inactivated serum. It is thought that EDTA could act by inhibiting a heat-labile component of human serum.     

Impact of extreme donor age on the outcome of living-related donor kidney transplantation

Eosinophil cationic protein (ECP) in sputum may be used to estimate the severity of bronchial inflammation and obstruction in asthmatics as well as to monitor asthma drug therapy. For this purpose, standardized processing of sputum is important. The aim of our study was to determine whether time and temperature influence the ECP concentration in the sputum of asthmatics. The samples of induced sputum obtained from 12 patients with stable asthma were homogenized using ultrasonification, and centrifuged. Supernatants were evenly divided and stored for 1, 6, 24 or 72 h at either 4 or 25 degrees C, then frozen at -80 degrees C. The ECP concentrations were determined using fluoroimmunoassay and compared with the immediately frozen samples. After storing at 4 degrees C, the ECP levels at the four time points were 101.2, 96.0, 98.2 and 90.6% of the initial concentration, respectively. When sputum specimens were stored at 25 degrees C, ECP levels decreased to 96.1, 94.4, 90.7 and 87.7%, respectively. The influence of time on ECP concentrations in sputa was statistically significant (p=0.02). A significant temperature effect was found when comparing the specimens stored at 4 degrees C with those at 25 degrees C (p=0.03). Looking at individual time points, a significant decrease in ECP concentration was only seen at 25 degrees C after 24 and 72 h. We conclude that eosinophilic cationic protein in the sputum of asthmatics decreases in a time- and temperature-dependent process. If sputa cannot be processed after obtaining the specimens, they should be stored in a refrigerator at 4 degrees C, until eosinophilic cationic protein is measured.     

Electron microscopy of postmortem autolysis of rat muscle tissue

To define the progression of ultrastructural changes in normal muscle at post mortem, rat gastrocnemius muscles were studied at various times after storage at +4 degrees C and +22 degrees C. Degeneration of the I-zone (discoid necrosis) and membranous bodies were found to be similar to that seen in muscle diseases, and lamellar formations were seen in mitochondria. At +4 degrees C there was contraction of the sarcomere which vanished in 12 h and inter-filamentous oedema appeared. Z-line degeneration was seen at 24 h and at 4 days all Z-lines had disappeared, and the H-zone showed darkening. In the same samples collapse or ruptures of the I band were seen. At 8 days the H-zones and M-lines were still discernible. In the early stages the mitochondria showed swelling and loss of matrix granules, while later they showed broken cristae and outer membranes, and flocculent densities. At 4 days rearrangement of the cristal material into long pentalaminar "needles" was seen in a few mitochondria. At 4 and 8 days membranous bodies were seen and the T-system and sarcoplasmic reticulum showed ruptures and disintegration into vesicles. The nucleus showed increasing condensation of chromatin at the periphery and clearing of the center. Polysomes and glycogen were reduced at 2 h, and has practically vanished at 24 h. At 22 degrees C the changes were the same but appeared about 4 times as quickly as at +4 degrees C.     

Subzero nonfreezing preservation in a murine limb replantation model

This study was designed to examine the most effective temperature for hypothermic storage, without freezing, to prolong ischemic tolerance in an amputated murine hindlimb model. We measured freezing points in the calf muscle and the subcutaneous tissue of the foot in the amputated limbs of Fisher 344 strain male inbred rats. The highest freezing point was -1.5 degrees C, which was recorded in the calf muscle. To prevent freezing in any of the tissues in the amputated limb, the temperature for the lowest nonfreezing preservation was defined as -1 degrees C. The amputated limbs were preserved at subzero nonfreezing temperature (-1 degrees C) and at 4 degrees C for 4, 8, 12, 24, 48, and 72 h, and were then transplanted to other inbred rats by microsurgical techniques. We evaluated the vascular patency of the anastomoses by direct observation and performed histological examinations on the seventh day after replantation. Subzero nonfreezing preservation of a limb at -1 degrees C for 72 h was significantly superior to hypothermic preservation at 4 degrees C for 72 h in terms of anastomotic patency rates (P < 0.05). The histology of skeletal muscles preserved at -1 degrees C for 8 h showed greater similarity to the normal situation than the histology of those preserved at 4 degrees C for 8 h. Bone viability with osteoblastic activity was maintained in grafted limbs preserved at -1 degrees C for 72 h, but in the limbs preserved at 4 degrees C for 72 h the bone was not viable, showing no osteoblastic activity. Clinically, the period of ischemia in major limb replantation at normal ambient temperatures is limited to about 6 h. In this study, the maximum ischemia time for replantation of a limb containing muscle tissue was prolong to 8 h at -1 degrees C, but the maximum ischemia time at 4 degrees C could not be prolonged to 8 h. Our results suggest that, in the major replantation of a limb containing muscular tissue, hypothermic preservation at -1 degrees C would be more useful than preservation at 4 degrees C.     

Effect of specimen storage on absolute CD4 counts

We have evaluated the effect of specimen storage on absolute CD4 counts by a commercially available manual assay. This assay utilizes latex particles coated with CD4 monoclonal antibodies that are mixed with lymphocytes in whole blood. Thirty blood samples were analyzed on days 1, 2, 4, and 7 postcollection. Linear regression analysis and Pearson correlation coefficients were used to determine the relationship between the absolute CD4 count and the storage time after sample collection. There was a significant decrease in absolute CD4 counts from baseline over time, dropping 3.6% at day 2, 10.1% at day 4, and 18.8% at day 7. However, the standard error of the B coefficient was constant [SE (B) = 0.031] up to day 4, indicating that reliable estimates of the baseline CD4 counts could be made from the CD4 counts determined up to day 4 from the time of sample collection. In addition to being sample, rapid, and inexpensive, the manual assay is capable of giving a reliable absolute CD4 count after specimen storage of up to 4 days. The application of this assay in the limited facilities of developing countries' laboratories is attractive.     

Zolpidem (Ambien): a pediatric case series

BACKGROUND: In 1993, the nonbenzodiazepine sedative-hypnotic zolpidem tartrate (Ambien) was approved for use in the US. Zolpidem has an imidazopyridine structure and possesses a rapid onset of action and a short half-life. The toxic threshold and profile have not been well established in the pediatric population. METHODS: All pediatric zolpidem exposures reported to a regional poison information center over 24 months were reviewed retrospectively from the American Association of Poison Control Centers Toxic Exposure Surveillance System data collection forms. RESULTS: Twelve pediatric zolpidem exposures were reported. Seven were unintentional (ages 20 mon-5 y) and five were intentional misuse/suicide (ages 12-16 y). The regional poison information center was contacted within 1 h in ten cases with onset of symptoms within 10 to 60 min (mean 31.6 min). One child had no effect with 2.5 mg. As little as 5 mg caused symptoms with minor outcome in six unintentional ingestions (5-30 mg). Minor to moderate symptoms were reported 1-4 h after intentional ingestions (12.5-150 mg). The duration of symptoms in the unintentional cases ranged from less than 60 min up to 4 h (mean 2.4 h) and 6-10 h (mean 7.5 h) in the intentional exposures. Treatment consisted of observation (4), syrup of ipecac (1), lavage and activated charcoal (1), activated charcoal alone (5), and unknown (1). CONCLUSION: Due to the very rapid onset of central nervous system symptoms in children, emesis is not a treatment option. Supportive care, activated charcoal in large ingestions, and observation until symptoms resolve may be sufficient in most pediatric cases.     

A gene essential to brain growth and development maps to the distal arm of rat chromosome 12

Studies were undertaken to determine the stability of nitrobenzodiazepines and their 7-amino metabolites in water and blood. At 22 degrees C nitrazepam and clonazepam were stable in sterile fresh blood containing preservative over 28 days, whereas 25% of flunitrazepam was degraded. At 37 degrees C all three drugs were substantially lost over 9 h (29-51%). There was only a small loss observed for the 7-amino metabolites and no substantial amounts of parent drug and 7-amino metabolite were degraded in water under these conditions. In the absence of preservative substantial amounts (25-50%) of parent drugs were lost in fresh blood over 10 days at 22 degrees C. In bacterially-contaminated postmortem blood all three drugs were completely degraded over 8 h at 22 degrees C with almost all drug completely converted to the respective 7-amino metabolite. These metabolites were also partially degraded (10-20%) over 45 h at 22 degrees C. All 3 nitrobenzodiazepines were stable in blood stored for up to 24 months at -20 degrees C, or 4 degrees C over 10 months. Their respective 7-amino metabolites were, however, relatively unstable at -20 degrees C with a significant loss (29%) after 2 months. At 4 degrees C a 21% loss occurred after 1 month. Freeze/thawing was found not to affect the concentration of nitrobenzodiazepine and 7-amino metabolites. These results show that the nitrobenzodiazepines and their metabolites are unstable chemically and metabolically in blood. We advise that blood collected for the purpose of nitrobenzodiazepine determinations should be preserved with sodium fluoride, stored at -20 degrees C and assayed as soon as practicable, preferably within a week of collection.     

Impact of various red cell concentrate preparation methods on the efficiency of prestorage white cell filtration and on red cells during storage for 42 days

BACKGROUND: White cell (WBC) reduction prior to storage of red cell (RBC) concentrates may reduce the incidence of HLA alloimmunization and may improve the quality of stored RBCs. STUDY DESIGN AND METHODS: An integrated WBC-reduction filter system was tested after various RBC preparation procedures (from whole blood), and the influence of filtration on RBCs during storage for 42 days was investigated. Four additive system RBC preparation protocols were used. Units prepared from conventional triple blood bags were held for 4 to 6 hours at 22 degrees C, and then the RBCs were separated via a hard spin and filtration, performed immediately (Group 1) or after 18 hours' storage at 4 degrees C (Group 2). Units prepared from a top-and-bottom collection system were held at 22 degrees C for 4 to 6 or 22 to 24 hours; the centrifuged RBCs were filtered immediately after preparation (Groups 3 and 4, respectively, by holding time). WBC reduction and filtration time were analyzed. The impact of WBC filtration on pH, hemolysis rate, hemoglobin content, ATP, potassium glucose, and lactate was investigated weekly during storage for 42 days. RESULTS: Filtration reduced the mean WBC count by 3 to 4 log10, to 0.19 +/- 0.25 x 10(6), regardless of the RBC preparation method. Mean filtration times differed significantly between the groups and were longest for Group 1. Besides hemolysis and pH values, which were greater in all filtered units, no major differences were found in filtered and unfiltered RBCs during the storage interval. CONCLUSION: The efficiency of prestorage WBC filtration of RBCs was unaffected by the preparation procedure. However, the filtration time for RBCs freshly prepared in the conventional triple blood bag system without buffy-coat depletion was unacceptable. No major metabolic differences between filtered and unfiltered RBCs during 42 days of storage were found.     

Mortality studies of machining fluid exposure in the automobile industry. IV: A case-control study of lung cancer

The taxanes represent a new class of clinical chemotherapeutic agents. A series of in vitro studies were independently of each other initiated in two different institutes (Amsterdam and Madison) to test the hypothesis that hyperthermia might enhance the cytotoxicity of taxanes. Clonogenic capacity experiments (Amsterdam) included the exposure of R1- and SW 1573-cells to 1, 4, or 24 h of paclitaxel with heat 43 degrees C x 60 min in the last hour of drug treatment or at 24, 48 as well as 72 h post drug treatment. Survival assay experiments (Madison) included the exposure of L-929-cells to paclitaxel and docetaxel for 24 h with heat 41.8 degrees C x 60 min the first or last hour of drug treatment as well as 24 and 48 h post treatment. No thermal enhancement of cytotoxicity for the taxanes was observed in these human and murine cell lines, with congruent data in both institutes. In addition, high performance liquid chromatography studies at 41.8 degrees C and 43 degrees C demonstrated paclitaxel and docetaxel were heat stable.     

Rapid determination of the membrane potential of mitochondria in small biopsy specimens of the liver

In liver transplantation, graft viability is ideally to be determined before implantation. Integrity of mitochondria may be a prerequisite to a viable graft. A new method is presented, which allows for the determination of the membrane potential of mitochondria (MPM; mV) in state 4 respiration within 50 min in 40-mg specimens, employing rhodamine 123 as a probe. Normal control showed a MPM of 239.2 mV. Storage in saline at 37 degrees C yielded an impaired MPM of 153.5 mV within 3 h. The cold storage at 1 degree C could preserve MPM at quasi-normal after 3 h but reduced it significantly after 24 h to 222.2 mV in saline (p < 0.005 vs. control) and 231.0 mV in UW solution (p < 0.05 vs. control): the difference between the 24-hour values was significant (p < 0.05).     

The survival and growth of acid-adapted mesophilic pathogens that contaminate meat after lactic acid decontamination

Lactic acid decontamination (LAD) may adapt pathogens to lactic acid. Such organisms may have an increased resistance to acid and can contaminate meat after LAD. The survival and growth of acid adapted Campylobacter jejuni, Salmonella typhimurium. Escherichia coli O157:H7 and Staphylococcus aureus inoculated on skin surface of still warm pork belly cuts 2 h after LAD was examined during chilled (4 degrees C) storage and refrigeration abuse equivalent to 12.5 degrees C. Lactic acid decontamination included dipping in 1, 2 or 5% lactic acid solutions at 55 degrees C for 120 s. Lactic acid decontamination brought sharp reductions in meat surface pH, but these recovered with time after LAD at approximately 1-1.5 pH units below that of water-treated controls. A sharp decrease in the number of cfu of pathogens occurred on chilled 2-5% lactic acid treated pork belly cuts when the skin surface was less than pH 4.8-5.2. The reductions ranged from 0.1-0.3 log10 cfu cm-2 for E. coli O157:H7 to over 1.7-2.4 log10 cfu cm-2 for Camp. jejuni, respectively. Increase in storage temperature from 4 to 12.5 degrees C reduced delayed decrease in numbers of all pathogens except Camp. jejuni by a factor of two. Deaths in Camp. jejuni at 12.5 degrees C slightly exceeded those at 4 degrees C. After the initial sharp decline, the number of cfu of mesophilic pathogens decreased gradually at a rate similar to that on water-treated controls. Growth of all mesophilic pathogens except Camp. jejuni on 2-5% LAD meat occurred during storage at 12.5 degrees C when the meat surface pH exceeded 4.8-5.2, and was slower than on water-treated controls. Low temperature and acid-adapted E. coli O157:H7, Salm. typhimurium and Staph. aureus, and acid adapted Camp. jejuni that contaminate skin surface after hot 2-5% LAD, did not cause an increased health hazard, although microbiota and intrinsic parameters (lactic acid content, pH) were created that could advantage their survival and growth.