51Cr Method Research Articles

Serial blood volume estimation in rabbits using trivalent chromium – An exploratory study

Though often used in cardiac intensive monitoring set up, simultaneous evaluation of several variables like CVP, PCWP, SVV and hTEE for fluid volume resuscitation (especially when capillary permeability is major problem than cardiac performance) is a major challenge in many ICU setups. Therefore, repetitive determination of blood volume by trivalent chromium [51Cr (III)] as a direct single variable method may be a near ideal method during fluid volume resuscitation in cases where capillary permeability is a major problem (e.g. Burns). Hence, in the present article the repeatability and reliability of 51Cr (III) method in New Zealand white rabbits was explored. Mean blood volume values of initial measurement were 195.66 ± 47.30 ml or 89.81 ± 17.88 ml/kg body weight. Repeated mean blood volume values, measured after 1 h, was 181.98 ± 53.16 ml or 83.68 ± 22.09 ml/kg body weight. The average difference between the initial and repeated measurements was 10.93 ml (95% CI −3.33, 25.19 ml), which is not statistically significant (P = 0.128). The method using 51Cr (III) for repeat blood volume measurements after sixty minutes in rabbits is a reliable method.•Rapid•Repeatable•Reliable

Posttransfusion red blood cell (RBC) survival determined using biotin‐labeled RBCs has distinct advantages over labeling with 51Cr

Posttransfusion red blood cell (RBC) survival determined using biotin‐labeled RBCs has distinct advantages over labeling with ⁵¹Cr

Red blood cell (RBC) volume can be independently determined in vivo in humans using RBCs labeled at different densities of biotin

Anemia is a serious problem in critically ill neonates. To investigate the pathophysiology of anemia and responses to red blood cell (RBC) transfusions and erythropoietin therapy, repeated measurement of red blood cell volume (RCV) and blood volume is useful. To extend our previous sheep study in which RBCs were labeled at four different biotin densities, we assessed the validity of this multidensity method for in vivo measurement of circulating RCV in humans. In eight healthy adults, autologous RBCs were biotinylated at each of four biotin densities (6, 18, 54, and 162 µg biotinylation reagent/mL RBC), mixed, and infused intravenously; blood was sampled at 10, 20, and 60 minutes. At each time, RCV was calculated from dilution of individual RBC populations enumerated by flow cytometry. RCV measurements from the population of RBCs biotinylated at 6 µg/mL were chosen as the reference values because this density had been previously validated against the 51Cr method in vitro and in vivo in humans. Values for RCVs were not significantly different among the four densities of biotinylated RBCs at any of the three time points and did not change over 60 minutes. These studies provide evidence that four densities of biotinylated RBCs can be used in vivo for simultaneous, independent, accurate measurements of RCV in humans. We speculate that this method will also be useful for repeated measurement of RCV and blood volume in infants and other patient populations in whom radioactive labels should be avoided.

Blood Loss During Transurethral Resection of the Prostate as Measured by the Chromium-51 Method

To evaluate blood loss during transurethral resection of the prostate (TURP), and its predictive factors, using the chromium 51 (51Cr) labeling method. From January to June 2008, 41 patients who underwent TURP for symptomatic benign prostatic hyperplasia (BPH) at four French urology centers were included in the analysis. Red cells volume was measured by the 51Cr method 1 day before TURP, and on postoperative day 3. Overall blood loss was estimated by multiplication of red cells volume loss and preoperative venous hematocrit value. Mean preoperative red cells volume was 1997 mL. Mean loss of red cells volume was 209 ml, which corresponds to an estimated blood loss of 507 mL. Mean delta of hematocrit and hemoglobin were 1.4% and 0.71 g/dL, respectively. In univariate analysis, prostate volume, weight of resected tissue, preoperative red cells volume, and resection time were significantly and directly associated with loss of red cells volume (P = 0.038, P = 0.004, P = 0.002, and P = 0.039, respectively). Bipolar and monopolar TURP did not lead to significant difference of red cells loss. In multivariate analysis, both preoperative red cells volume and weight of resected tissue were independent predictors of red cells loss (P = 0.017 and P = 0.048 respectively). We present the first study to measure blood loss secondary to TURP using the 51Cr method. This technique allowed evaluating blood loss not only during the surgical procedure but also during the postoperative period. We learned from this study that, on average, blood loss from the procedure until postoperative day 3 was more than 500 mL, which is larger than previously reported amounts as measured by other methods. Because significant blood loss might occur during the postoperative period, the 51Cr method should be used to measure blood loss when evaluating new emerging techniques to manage BPH.

Prediction of red cell and blood volumes distribution by various nomograms: do current nomograms overestimate?

Currently used formulas for estimation of a person's red cell volume (RCV) by weight and height are decades old and were based on the use of 51Cr isotopes and on a sample population, which may not be reflective of today's population. In this study, the accuracy and precision of the use of 99mTc RCV measurements in volunteers more typical of today's population were evaluated. The subjects were volunteers who met the requirements for a standard blood donation. The mean +/- standard deviation (SD) 99mTc RCV for 127 males (mean weight, 83.2 kg; height, 180 cm) was 2062 +/- 339 mL, and for 101 females (mean weight, 69.5 kg; height, 166 cm) it was 1320 +/- 201 mL. These results were highly correlated with RCV results with the standard extrapolation 51Cr method with stored red blood cells (RBCs) and highly consistent (within +/-10%) by repeated measurements with the same 22 donors over a 3.5-year period. The RCV results correlated with estimates from the current formulas, but were on average 11 to 14 percent lower. The studies demonstrated that 99mTc is a reproducible and precise method for determination of a person's RCV and that current formulas may significantly overestimate the RCV of today's population. This is likely the result of a shift in population characteristics over the past four decades as reflected by an increased mean body mass index (from 25 to 28 kg/m2), which has not resulted in a proportionally increased RCV.

Hyperhemolysis during the evolution of uncomplicated acute painful episodes in patients with sickle cell anemia

Some patients with sickle cell anemia (SS) experience significant decrease in their hemoglobin (Hb) level and significant increase in reticulocyte count during the progression of uncomplicated acute painful episodes. These changes have been attributed to, but not proven, hyperhemolysis. This study provides evidence of hyperhemolysis in this group of patients. Hospitalized patients who met the above criteria were identified and followed prospectively, and the measures of red blood cell (RBC) survival were determined. The latter included 1) laboratory measures of hemolysis: Hb level, reticulocyte count, nucleated RBC count, RBC distribution width (RDW), total bilirubin, lactate dehydrogenase (LDH), and aspartate transaminase (AST); 2) determination of RBC survival in vivo by the (51)Cr method in selected patients; and 3) the determination of the RBC Hb:reticulocyte Hb ratio. Baseline values of these measures were also obtained and compared to the crisis data. During the evolution of the painful episode in the selected patients, there was a significant increase in total bilirubin, LDH, AST, nucleated RBC count, and RDW that are indicative of hyperhemolysis. There was a significant decrease in RBC Hb:reticulocyte Hb and (51)Cr RBC survival, which is diagnostic of hyperhemolysis. Together, the data indicate that hyperhemolysis occurs during uncomplicated acute painful episodes in some patients with SS. The presence of free plasma Hb, consequent to hyperhemolysis, reduces nitric oxide bioavailability, promotes endothelial dysfunction, and contributes to the development of pulmonary hypertension and vasoocclusion.

Accuracy of carboxyhemoglobin dilution method for the measurement of circulating blood volume.

The management of circulating blood volume (BVc) is crucial in intensive care unit (ICU) patients. The purpose of this study was to verify the accuracy and precision of the carbon monoxide-labeled hemoglobin (CO-Hb) dilution method (CO method) by comparing it with the 51Cr-labeled erythrocyte dilution method (51Cr method) for the measurement of BVc. A prospective study was performed in 18 patients who underwent coronary artery bypass grafting (CABG) under mild hypothermic cardiopulmonary bypass (CPB). The BVc was measured by both the CO method and the 51Cr method at 24 hr after ICU admission in order to verify the accuracy and precision of the CO method. Paired data were assessed in absolute terms, and percentage errors were calculated by the degree of agreement. Small mean differences and standard deviations between the CO method and the 51Cr method (-70.2 +/-184.8 mL) and small percentage errors (-0.49+/-1.29%) indicated the accuracy and precision of the CO method, and a close correlation was observed (r = 0.97). The CO method can measure BVc with a similar degree of accuracy as the 51Cr method. It is simple, repeatable and safe without the risk of exposure to radioactivity in the ICU.

Determinants of Red Cell Survival and Erythropoietic Activity in Patients with Sickle Cell Anemia in the Steady State

We have studied 26 patients with sickle cell anemia to determine the factors that affect red blood cell (RBC) survival and other parameters of erythropoietic activity in the steady state. Determinants of erythropoietic activity included RBC survival by the 51Cr method, RBC production/destruction rate, a genotype, βs haplotype, plasma 59Fe clearance, plasma iron turnover, eryth-ron transferrin uptake), RBC Fe utilization, reticulocyte count, and erythro-poietin levels. The α genotype was the most significant determinant of RBC survival followed, to a lesser extent, by the βs haplotype. Hb F showed no correlation with RBC survival due to patient selection bias - the patients studied had comparable Hb F levels to start with. Other determinants of erythropoietic activity (hemoglobin level, mean corpuscular volume, reticulocyte count, RBC mass, RBC production/destruction rate, and erythropoietin level) were most likely secondary determinants associated with the α genotype, and not independent determinants in themselves. The data suggest that the α genotype and, and to a lesser extent, the βs haplotype, might be determinants of the severity of the anemia of sickle cell disease, and should be considered in genetic counseling and patient selection for aggressive therapeutic interventions.

Erythropoietic activity in patients with sickle cell anaemia before and after treatment with hydroxyurea

In this project we have prospectively studied the erythropoietic activity in patients with sickle cell anaemia (SS) before and after treatment with hydroxyurea (HU). Some of the patients were enrolled in a double-blind placebo controlled trial of HU in patients with SS and others were enrolled in an open label study. Determinants of erythropoietic activity included the reticulocyte count, red blood cell (RBC) survival by the 51Cr method, plasma 59Fe clearance, plasma iron turnover (PIT), erythron transferrin uptake (ETU), RBC production/destruction rate, and RBC Fe utilization. Therapy with HU increased the mean corpuscular volume (MCV), haemoglobin (Hb)F, RBC survival and t1/2 59Fe clearance; it decreased the reticulocyte count, the white blood cell (WBC) count, ETU, and the PIT. Most of the changes in parameters of erythropoiesis could be explained by the increase in 51Cr RBC survival after therapy with HU. Together the data showed that in selected patients the net effect of HU on Hb level was a function of the difference between the suppressive effect of HU (decreased RBC production) and the increase in RBC survival. In the majority of patients who responded to HU, there was a preferential effect on RBC survival.

High predictive value of red cell volume measurement using carboxy-haemoglobin in a rabbit model of haemorrhage

We have studied the accuracy of blood volume measurements using carbon monoxide (CO)-labelled haemoglobin (COHb) injection and dilution (CO method) by comparing changes in red cell volume (RCV) measured using the CO method and 51Cr-labelled erythrocyte dilution (51Cr method) in a haemorrhage and infusion model in rabbits. RCV was measured repeatedly using the CO method at four different blood volume stages (stages I-IV). At stages I and IV, RCV was measured simultaneously using the 51Cr method. In comparing the sum of the circulating RCV and extracted RCV (SUM RCV) using the CO method, the values were almost equal and there were no significant differences between the values at the four stages. In comparing circulating RCV measured using the CO method and the 51Cr method, mean difference between the two methods was 0.80 (SD 0.76) ml kg-1 or 4.7 (4.6)%, and a positive correlation was observed (r = 0.91). We conclude that the CO method can be used to measure blood volume during perioperative periods in infants because it avoids use of a radioactive tracer, is simple and repeated measurements are possible.

Comparison of tetrazolium colorimetric and 51Cr release assays for cytotoxicity determination of dental biomaterials

Objectives. The purpose of this study was to compare a methylthiazole tetrazolium (MTT) dye colorimetric method with the standard 51Cr assay as methods of assessing cytotoxicity of dental materials. Methods. Two MTT-based colorimetric formats, test tube and 96-well microplate methods, were compared to the 51Cr release assay. A series of eigth dental materials were evaluated. Cytotoxicity profiles were determined for each test material. A TC 50 value (Toxic Concentration required to kill 50% of the cells) was determined for each biomaterial, and these results were used to make statistical comparisons between the methods. Results. The three methods were statistically correlated (p < 0.005) by comparison of the eight samples tested. That is, the same rank in toxicity was given by the two tetrazolium sample formats and the 51Cr method. Significance. The MTT assay was found to have several advantages in comparison to the current standard 51Cr release assay. Optimized in the 96 well format, complete dose response curves and greater sample comparisons can be made rapidly, making the MTT method more economical in time and cost. Furthermore, the MTT method is based on intracellular biochemical changes, measuring cell viability rather than cell morbidity, and has lower detectable limits than the 51Cr release method. There is also less detector chemical binding interference than encountered in the 51Cr release method.

Introduction of a Well-type germanium/lithium semiconductor detector in nuclear medicine

Semi-in-vivo diagnostic procedures in nuclear medicine are commonly carried out using conventional scintillation detectors. Modern semiconductor detectors have rarely been applied in this field until now. A particular example of this group is the Well-type germanium/lithium detector, a semiconductor detector which combines a high energy resolution with a high efficiency for gamma-radiation. The latter, combined with an excellent signal-to-background ratio, allows accurate detection of even very low activities. Patient doses can therefore be reduced considerably, as was demonstrated in erythrocyte survival studies, platelet survival studies and the Schilling test. The effective dose equivalent associated with erythrocyte survival studies, using the recommended 51Cr method, could be reduced from 780 to 2 μSv. The effective dose equivalent in platelet survival studies, using the recommended 111In method, could be reduced from 1400 to 14 μSv. For both procedures, the reliability was demonstrated by the results obtained with healthy controls. In the Schilling test, using 57Co, the effective dose equivalent could be reduced from 54 to 9 μSv. In addition, the high resolution of Ge-detectors enables selective and accurate determination of individual components in mixtures of isotopes. Therefore, with such detectors, low-level double-labeling procedures can be introduced in semi-in-vivo nuclear medicine, thus creating the possibility of studying certain unanswered but clinically relevant questions.

Response of rainbow trout to constant-pressure and constant-volume hemorrhage.

Plasma oncotic pressure (OC), protein concentration (P), osmolality (Osm), and hematocrit (Hct) were measured in conscious rainbow trout subjected to 120 min of constant-pressure (CPH) or constant-volume (CVH) hemorrhage. Blood volume, tissue red cell, and plasma space were determined in control and CPH trout with 51Cr-labeled red cells and 125I-albumin. At the onset of CPH, blood loss exceeded 1.5 ml.min-1.kg-1 but quickly dropped to a constant rate of 0.1 ml.min-1.kg-1 by 15 min. After 2 h, 73% of the initial blood volume was lost, and Hct, OC, and P were reduced by 55 +/- 3, 39 +/- 4, and 19 +/- 9%, respectively; Osm did not change. Calculated blood volume was reduced by 39% (51Cr method) or by 56% (125I method). Red cell space was reduced in all tissues except brain. Plasma space decreased in stomach, increased in liver and kidney, and was unchanged in other tissues. CVH had similar effects on Hct, OC, and P, while Osm and plasma sodium increased. The time required for arterial pressure to recover to within 90% of control increased with successive bleeding from 39 to greater than 160 min. Twenty-four hours after CVH, Hct remained depressed, while all other parameters were returning toward control levels. Trout incorporate a variety of mechanisms to respond to hemorrhage, including mobilization of stored blood and reabsorption of fluid from interstitial and probably intracellular sources. High permeability of trout capillaries to protein may facilitate the restoration of protein and fluid and allow fish to survive extensive hypovolemia.

Determination of single and repeated red cell volumes by the indicator dilution method using carbon monoxide as the indicator.

The use of radioactive isotopes limits clinical applications of blood volume measurement in the ICU. We measured red cell volumes with carbon monoxide-labeled RBC in six dogs and five human volunteers. The measured values obtained on the dogs were compared with the simultaneous measurements with the 51Cr method; the ratio of the carbon monoxide to 51Cr values ranged from 0.86 to 1.17, and the mean ratio was 1.0 +/- 0.1 (SD), r = .93. We infer from these results that the carbon monoxide method has several advantages over the 51Cr method: a) the short labeling time (about 1 min), b) rapidly decreasing background levels of carbon monoxide with FIO2 1.0, and c) repeatability at intervals of several hours.

An improved method for the in vivo labelling of red blood cells with 99mTc in gated cardiac imaging.

Red blood cells of 10 patients were labelled with 99mTc by the in vivo method, 5 of the patients were orally administered with 400 mg potassium perchlorate at the time of stannous pyrophosphate injection and 5 ml blood was withdrawn at 10 min postinjection of pertechnetate to determine the labelling efficiency of RBC's. The RBC and plasma volumes were also determined in all patients by the 51Cr method. Our results indicated higher incorporation of injected radioactivity into RBC's in the perchlorate administered group (83.8% +/- 9.2%) compared to the other (58.5% +/- 22.0%) (P less than 0.01). The thyroid and stomach were visualized in the control group, but not in the perchlorate administered group. In routine application, 50 MUGA studies have been done with perchlorate administration with excellent results. In 60 patients without perchlorate administration previous to the present study the EF could not be determined in 6 patients (10%). Our results indicated that potassium perchlorate may be used routinely for in vivo labelling of RBC's to improve the labelling efficiency and the quality of gated cardiac imaging.

Gastrointestinal Blood Loss in Arthritic Patients Receiving Chronic Dosing with Etodolac and Piroxicam

Etodolac, a new anti-inflammatory analgesic drug found to be effective in treating arthritis in a dose range of 100 to 300 mg bid, has been shown to induce significantly less gastrointestinal microbleeding in normal men than several other NSAIDs. In this study, the effect on gastrointestinal blood loss of high-dose etodolac, 300 and 500 mg bid, versus piroxicam at its normal therapeutic dose of 20 mg qd, was investigated by the 51Cr method in 23 men with osteo- or rheumatoid arthritis. Placebo periods preceded and followed 28 days of active drug treatment. Blood and stool analyses were performed by an analyst not aware of drug assignment or study design. Patients receiving piroxicam, but not those receiving either dose of etodolac, had a significantly higher mean level of fecal blood loss in the active treatment phase compared with the pretreatment placebo level (p less than 0.01). Further, microbleeding was significantly greater for the piroxicam group during treatment than for either of the etodolac groups (p less than 0.01). There were no significant differences in fecal blood loss between the two groups receiving etodolac compared with pretreatment. Even at doses two to three times those found effective in the treatment of arthritis, etodolac produces no increase in fecal blood loss, in contrast to blood loss seen with the recommended dose of piroxicam. Fecal blood loss in osteoarthritic patients, not receiving an NSAID, was similar to normal subjects in previous studies.

Flow cytometric and radioisotopic determinations of platelet survival time in normal cats and feline leukemia virus-infected cats.

This study demonstrates the potential usefulness of a flow cytometric technique to measure platelet survival time in cats utilizing autologous platelets labeled in vitro with fluorescein isothiocyanate (FITC). When compared with a 51Cr method, no significant differences in estimated survival times were found. Both the 51Cr and FITC-labeling procedures induced similar changes in platelet shape and collagen-induced aggregation. Platelets labeled with FITC had significantly greater volumes compared with those of glutaraldehyde-fixed platelets. These changes were primarily related to the platelet centrifugation and washing procedures rather than the labels themselves. This novel technique potentially has wide applicability to cell circulation time studies as flow cytometry equipment becomes more readily available. Problems with the technique are discussed. In a preliminary study of the platelet survival time in feline leukemia virus (FeLV)-infected cats, two of three cats had significantly reduced survival times using both flow cytometric and radioisotopic methods. These data suggest increased platelet turnover in FeLV-infected cats.

Estimation of transfused red cell survival using an enzyme‐linked antiglobulin test

An enzyme-linked antiglobulin test (ELAT) method was developed to estimate survival of transfused red cells. This procedure is based on a principle analogous to that of the Ashby technique were antigenically distinct red cells are transfused and their survival studied. We compared the ELAT survival to the 51Chromium method (51Cr) in four patients. Three patients with hypoproliferative anemias showed T 1/2 by ELAT of 17.5, 18, and 17 days versus 18.5, 20, and 19 days by the 51Cr method. A fourth patient with traumatic cardiac hemolysis had two studies performed. In this case, the ELAT showed a T 1/2 of 10 and 8.1 days while 51Cr T 1/2 values were 11 and 10.5 days. The ELAT method for measuring red cell survival yielded data which agreed closely with the results of the 51Cr method. Although 51Cr is the accepted method for red cell survival, the ELAT method can be used to estimate transfused red cell survival.

Use of 75Se-labeled methionine to study the sequestration of senescent red blood cells.

Labeling red blood cells with Na251CrO4 enabled us to study certain aspects of red cell survival and sequestration from the circulation. As a random labeling procedure, however, the 51Cr method has certain limitations. Therefore, we developed a cohort labeling method using 75Se-methionine as a two-rat procedure. This gives us a clear pulse-labeled population of rat red cells to study the dynamics of sequestration. With this labeling procedure, it was possible to demonstrate that 1) there is an increase in the density of red cells with age, 2) a significant sequestration of red cells from the circulation is apparent at the end of 48 days and essentially is complete at the end of 60 days, 3) there is a corresponding uptake of senescent red cells in the spleen, which peaks at 55 days, and 4) the 60-day end point is sharper and is more definitive when the "specific activity" (cpm per red blood cell) of the labeled red cells in the spleen is compared to that of the red cells still in the circulation. Asialo red cells, obtained by removal of sialic acid with sialidase, frequently have been used as a model for the study of sequestration of senescent red cells. With the technique herein described, it was possible to show that while asialo red cells will inhibit the uptake of labeled asialo red cells, they have no effect on the sequestration of senescent red cells. Presumably, different sites and mechanisms of sequestration are involved.

Occult Faecal Blood Loss Determined by a51Cr Method and Chemical Tests in Patients Referred for Colonoscopy

In 67 patients referred for colonoscopy the faecal blood loss was determined by a 51Cr method and 7 chemical tests. For patients with negative colonoscopy (no. = 10), colorectal polyps (no. = 24), rectal cancer (no. = 8), or colonic cancer (no. = 12), the median 51Cr-determined faecal blood loss was 0.67, 0.74, 1.26, and 2.18 ml/24 h, respectively. For all chemical tests the results were highly influenced by the upper time limit for a positive reaction. Mixing of faecal specimens before testing proved unimportant. Fecatwin sensitive showed more positive tests in delayed compared with immediate analyses (p less than 0.01). Of cases of colorectal polyps, tetramethylbenzidine (TMB) tests including Hemo-Fec Test could detect half, the benzidine test 2 of 5. Fecatwin sensitive and Hemoccult II 1 of 4, and Fecatwin 1 of 24. Of cases of colorectal cancer, TMB tests, the benzidine test, Fecatwin sensitive, Hemoccult, and Fecatwin could detect about 85%, 85%, 85%, 80%, and 45%, respectively. All chemical tests detected faecal blood loss from colorectal lesions more easily than from gastric lesions.