A brief history of hemoglobinometry.
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The surveys of the College of American Pathologists indicate a consistent bias between the manual "reference" method for hemoglobin determinations and the Coulter S measurements. The Coulter S hemoglobin values are invariably lower; the difference averages 0.3 g/dl. The commonly used calibration methods for the Coulter appear to be subobtimal. The hemoglobin calibration and quality assurance method proposed by Bull and colleagues is advocated as the presently most acceptable method. Commercial hemoglobin control materials should not be used for hemoglobin calibration.
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Properties of a quality control material for blood gases and pH should be similar to normal human whole blood with respect to oxygen buffering and acid-base behaviour. A hemoglobin solution may potentially fulfill this. However, the drawbacks of such a solution are the high oxygen affinity (lowp50), especially when it is prepared from human blood, and the improper concentration of bicarbonate. Bicarbonate is added to human stroma-free hemoglobin solution (SFHS), prepared as described previously, to obtain the desired pH and pCO2 combinations. Tonometry was used to determine the appropriate concentration of bicarbonate, which is 22.5 mmol/L to obtain an acidotic, and 29 mmol/L for both an alkalotic, and normal pH and pCO2 combination. Inositolhexaphosphate (IHP) is added to SFHS containing bicarbonate to obtain a normal p50 (around 3.55 kPa). Tonometry was used to determine the molar ratio of IHP/Hb4 (mol/mol) at which this is achieved. The molar ratios of IHP/Hb4 are 1.52, 1.74 and 3.40 for preparations with an acidotic, normal and alkalotic pH, respectively. In human SFHS nHill is 2.55 in the absence of IHP,nHill is at minimum 1.71 at a molar ratio IHP/Hb4 of 1.86 and increases to 2.53 at a molar ratio IHP/Hb4 of 5.04 and higher. Because the p50 will decrease with chi Hi this was studied at molar ratios of IHP/Hb4 of 0, 2 and 4, which covers the range of ratios as used. At these molar ratios of 0, 2 and 4, the decrease in p50 is 0.017 kPa/%Hi, 0.023 kPa/%Hi and 0.028 kPa/%Hi, respectively. Because bovine Hb was p50 near that of normal human blood, it is also used. The oxygen affinity shows a small decrease (p50 increases from 3.05 to 5.27 kPa) on addition of IHP. In the absence of IHP, nHill is 2.51 and nHill is at maximum 3.35 at molar ratios IHP/Hb4 between 3.00 to 4.56. At higher molar ratios nHill decreases to 2.90.
Human blood haemolysates containing ethylene glycol (final volume fraction 0.35) were prepared and stored at -20 degrees C (in the liquid state) up to 372 days. During the whole period, the total haemoglobin concentration (assayed in the material by means of the reference HiCN method) was found to be stable; spectral analysis also failed in detecting any deterioration, Hi formation being low. Good stability was also recorded on storage at 2-4 degrees C for 15 days, but only for 1-2 days at room temperature. The stabilised haemolysate is suggested as a material for long-term control of accuracy in hemoglobinometry.
Actual circulating blood volume during dialysis therapy can be monitored by continuous hemoglobinometry. Using this method in 15 stable, clinically nonoverhydrated dialysis patients, blood volume was recorded applying different modes of ultrafiltration: constant ultrafiltration (less than 500 ml/hr); high initial (greater than 1,500-2,000 ml/hr), subsequently decreasing ultrafiltration; and intermittently high (greater than 1,500 ml/hr) ultrafiltration. Mean amount of ultrafiltrate in all patients was 3,400 ml. Mean decrease in blood volume by 20% was generally tolerated without a decrease in blood pressure. Irrespective of the different modes of ultrafiltration, a decrease in blood volume was dependent only on the amount of ultrafiltered fluid. A constant, low ultrafiltration rate was not superior to a high ultrafiltration rate. In stable dialysis patients, decrease in blood volume is dependent only on the amount of ultrafiltrate. Up to a 20% decrease in blood volume, fluid can be removed from the patient even at a rate of 2,000 ml/hr.
Symptomatic hypotension due to ultrafiltration (UF) is one of the most frequent unwanted side effects of dialysis therapy. Using hemoglobinometry for continuous monitoring of blood volume (BV), ultrafiltration rate (UFR) can be adapted to actual changes in BV. A control system is shown in which UFR is set according to a predefined profile of BV. The conditions of control are: relative BV shall decrease steadily; BV shall decrease rapidly during the first 60 min of dialysis, thereafter decrease in BV should be less; UFR shall be as high as possible; and dry weight should be obtained within a given time. Application of this controlled UF method in 10 dialysis patients shows significantly fewer hypotensive periods and muscle cramps compared to conventionally constant UFR. It is concluded that BV controlled UF is an important step toward optimizing dialysis therapy.
The correlations between actual blood volume (BV), blood pressure (BP), heart rate, and plasma levels of renin activity (PRA), serum aldosterone (ALD), antidiuretic hormone (ADH), epinephrine (E), norepinephrine (NE), atrial natriuretic factor (ANF), cGMP, and cAMP were investigated in 10 stable patients during HD. HD consisted of four periods of about 60 min each. One half with an UF rate greater than 1,000 ml/h, followed by a time interval of 30 min without UF resulting in a "saw tooth" profile of BV. Decrease in BV was measured by continuous hemoglobinometry. Average total decrease in BV was 25%, while BP and HR did not change significantly. E, NE, ANF and ADH levels were directly related to actual changes in BV, suggesting that BP regulation in this special mode of HD is mainly supported by endogenous catecholamine and ADH secretion. The second messenger cGMP did not follow actual BV changes, but showed a significant decrease correlated with diminished BV. A significant change in PRA and ALD was missing. It is concluded that vascular stability in these patients is maintained by the response of catecholamins and ADH to decrease in blood volume, and not by the renin-aldosterone system.
Hemoglobinometry according to the International Committee of Standardization in Hematology (ICSH) suffers from imprecision related to high sample dilution and from potential errors owing to sample turbidity. We have evaluated a new instrument, "HemoCue," that measures hemoglobin at two wavelengths as azide methemoglobin, without dilution. The HemoCue method is superior to the ICSH method: by correction for turbidity, it avoids false hemoglobin readings that may arise from hyperlipemia or some large M-component of the immunoglobulin M class. We find the equipment suitable for use in outpatient units.
In patients on chronic hemodialysis the prevalence of atherosclerosis is increased and is by far the leading cause of morbidity and mortality. Endothelin-1, an endothelium-derived peptide with vasoconstrictive and mitogenic effects on vascular smooth muscles, is involved in the pathogenesis of atherosclerosis. The aim of the present study was to investigate the time course of plasma endothelin-1 levels during a hemodialysis session and to explore the influence of preexisting type 2 diabetes mellitus. Forty-five clinically stable hemodialysis patients (21 females, 24 males; mean age 62 +/- 12 years) were evaluated. Patients with type 2 diabetes (n= 11) were compared with the group of patients without diabetes (n=34). Relative blood volume (BV) changes (hemoglobinometry) and blood pressure (BP) was measured. Samples were taken before, every hour during, and after hemodialysis. Plasma endothelin-1 levels were measured by enzyme-linked immunoassay (ELISA) and results were corrected according to hemoconcentration. Hemodialysis with an ultrafiltration of 2215 +/- 952 mL was performed. Total BV at the end of hemodialysis was 89.3% +/- 8.3% of the pretreatment volume. Plasma endothelin-1 was enhanced in hemodialysis patients compared to normal subjects and increased from 1.28 +/- 0.47 before to 1.44 +/- 0.54 pg/mL (ref. 0.3-0.9) at the end of hemodialysis (p<0.05). The BV change (r=0.41) and the BP (mean BP: r=0.34) correlated with plasma endothelin-1 at the end of hemodialysis (p<0.05). The levels of endothelin-1 were significantly higher in the group of dialysis patients with type 2 diabetes compared to nondiabetics in all measurements (p<0.05). These findings suggest a potential role of endothelin-1 in the pathogenesis of vascular dysfunction in diabetes mellitus. The dialysis procedure per se, through vasoconstriction due to BV decrease, local endothelial injury (a.v. fistula), or bioincompatibility reactions (foreign surface contact) may additionally alter endothelial cell functions.
In patients on chronic hemodialysis hypotensive episodes are frequently encountered during the course of treatment and the prevalence of atherosclerosis is increased. Endothelin-1 (ET-1), an endothelium-derived peptide with vasoconstrictive and mitogenic effects on smooth muscles, is involved in vascular tone regulation and in the pathogenesis of atherosclerosis. The aim of the present study was to investigate plasma ET-1 during hemodialysis treatment and to explore the probable influence of pre-existing hypertension. Forty-seven hemodialysis patients (21 females, mean age 62 +/- 12 years) were evaluated and hypertensive patients (n = 33) were compared to normotensive patients (n = 14). Relative blood volume changes (hemoglobinometry) and blood pressure were measured. Samples were taken before, every hour during and after hemodialysis. Plasma ET-1 was measured by enzyme-linked immunosorbent assay and results were corrected according to hemoconcentration. Hemodialysis with an ultrafiltration rate of 2224 +/- 933 mL was performed. Total blood volume at the end of hemodialysis was 89.4 +/- 8.2% of the pretreatment volume. The fall in blood pressure (137/74 +/- 22/11 mmHg vs 127/73 +/- 30/14 mmHg) correlated with the decrease in blood volume (mean blood pressure: r = 0.33). Plasma ET-1 increased from 1.29 +/- 0.47 pg/mL before to 1.46 +/- 0.56 pg/mL (reference range 0.3-0.9) at the end of hemodialysis (P < 0.05). This rise was more pronounced in patients with hypertension than in normotensive individuals (P < 0.05). The change in blood volume (r = 0.41) and blood pressure (mean blood pressure: r = 0.34) correlated with plasma ET-1 at the end of hemodialysis (P < 0.05). Plasma ET-1 was enhanced in hemodialysis patients compared to normal subjects. During the hemodialysis session an increase in ET-1 was encountered, which was more pronounced in hypertensive than in normotensive patients and paralleled the hemodynamic changes. Apart from pre-existing hypertension, further factors potentially influencing ET-1 include local endothelial injury (arteriovenous fistula) and generalized bioincompatibility reactions (e.g. foreign surface contact) occurring during hemodialysis.
Correlations of energy state with response to therapy are more difficult to analyze because of the large effect of tumor clearing and oxygenation upon the tumor energy state as detected by PMRS alone. The combination of time-resolved hemoglobinometry using picosecond laser technology and localized PMRS seems appropriate to unravel the complexities of therapeutic intervention, tumor energetics, and oxygenation.
The changes in blood volume (BV), atrial natriuretic peptide (ANP), plasma renin activity (PRA), aldosterone (Aldo), norepinephrine (NE), epinephrine (Epi), parathyroid hormone (PTH), arginine vasopressin (AVP) and the cyclic nucleotides cAMP and cGMP were measured during a fluctuating BV cycle in 15 patients with end-stage renal failure maintained on chronic hemodialysis (HD). HD consisted of 4 periods of about 60 min each. The first half of each HD period consisted of ultrafiltration (UF) greater than 1,000 ml/h, and the second half consisted of no UF. Changes in relative BV were measured using continuous hemoglobinometry. Total BV at the end of treatment was 74.3 +/- 6.9% of the pretreatment volume. A significant positive correlation between BV and the levels of ANP, PTH, Epi and cGMP and an inverse correlation between BV and PRA, Aldo, AVP and NE were demonstrated. While mean values of NE and AVP levels were directly related to actual changes in BV, individual values did not homogeneously reflect this relationship. The cyclic nucleotides cGMP and cAMP did not follow immediate BV changes, but showed a significant decrease correlated with diminished BV. Based on a pre-postdialysis analysis, significant changes in PRA and Aldo were missing. It seems possible that vascular stability in dialysis patients may be maintained by the response of NE and AVP, and not by the renin-aldosterone system. The changes in ANP and cGMP values correlated most significantly (r = 0.38 and r = 0.51, p < 0.005) with the changes in BV, but no single variable could explain the blood pressure regulation during HD with intermittent rapid UF.
BACKGROUND: Several effects of hemodialysis, including hemoconcentration, alterations of hemostasis or hemorheology and endothelial activation, could potentially interfere with cerebral blood flow (CBF) regulation. These treatment-specific changes may also be crucial for the enhanced incidence of stroke in uremic patients. Nevertheless, the influence of hemodialysis on CBF has not been yet adequately studied. PATIENTS AND METHODS: We registered mean blood flow velocity (MFV) in the middle cerebral artery (MCA) during hemodialysis treatment in order to evaluate its contribution on CBF changes. Transcranial Doppler ultrasonography (TCD) of the MCA was performed continuously during hemodialysis treatment in 18 stable patients (10 males and 8 females, mean age 62 +/- 11 years) with end-stage renal disease of various origin. Blood pressure (mmHg), heart rate (/min), ultrafiltration volume (ml), BV changes (deltaBV by hemoglobinometry, %), arterial blood gases (pO2, blood oxygen content, pCO2), hemostasis activation (thrombin-antithrombin III complex, ELISA) and fibrinogen (Clauss) were measured simultaneously at the beginning of treatment and every hour thereafter. RESULTS: Before the hemodialysis session the MFV in the MCA was within normal range (57.5 +/- 13.0 cm/s, ref. 60 +/- 12) and was mainly dependent on the patients' age (r = -0.697, p < 0.01). The blood flow velocity in the MCA decreased significantly from 57.5 +/- 13.0 cm/s before the beginning to 48.3 +/- 11.1 cm/s after four hours (n = 18, p < 0.05) and to 43.9 +/- 8.9 cm/s after five hours (n = 9, p < 0.05) of hemodialysis treatment. During hemodialysis treatment, the percentual changes of MFV in the MCA (delta%MFV) were interrelated to the ultrafiltration volume (r = -0.486, p < 0.01), the blood volume (BV%, r = 0.369, p < 0.01) and the percentual changes of the hematocrit (r = -0.358, p < 0.01), of the arterial blood oxygen content (delta%acO2, r = -0.420, p < 0.01) and of the plasma fibrinogen levels (delta%fibrinogen, r = 0.244, p < 0.05). CONCLUSION: A significant continuous decrease of the MFV in the MCA was observed during hemodialysis treatment, which inversely correlated both with ultrafiltration volume, BV changes and changes of plasma fibrinogen. The ultrafiltration-induced hemoconcentration with concomitant rise of hematocrit and oxygen transport capacity, may partly explain the alterations in the cerebral MFV observed during hemodialysis.
The correction of anemia with human recombinant erythropoietin (rHuEPO) in end stage renal disease is associated with hypertension in about one third of hemodialysis patients. The pathogenesis of the rHuEPO-induced hypertension is still uncertain, though evidence of the involvement of endothelial cells has emerged. The aim of this study was to determine plasma endothelin-1 during hemodialysis and to compare the endothelin-1 levels in hemodialysis patients with and without rHuEPO substitution. Nineteen stable patients (13 male and 6 female, mean age 62 +/- 11 years) with end stage renal disease were studied. Cuprophan dialysers (GFS 12, Gambro, Lund, Sweden) were used for hemodialysis in all cases. rHuEPO (40 U/kg s.c.) was administered to 10 patients. Blood pressure (BP; RR mmHg) and blood volume changes (deltaBV; hemoglobinometry %) were serially measured. Samples were taken before and every hour during hemodialysis. Plasma endothelin-1 was measured by ELISA (R&D Systems, Minneapolis, USA) and corrected for hemoconcentration. Endothelin-1 concentration was elevated before commencement of hemodialysis (1.16 +/- 0.36 pg/ml) when compared to healthy controls (ref. 0.3-0.9) and increased to 1.47 +/- 0.51 pg/ml by the end of the session (p<0.05). In patients under rHuEPO-substitution plasma endothelin-1 was higher when compared to patients without substitution before (1.25 +/- 0.3 vs. 1.05 +/- 0.3 pg/ml) and at the end of HD (1.62 +/- 0.5 vs. 1.28 +/- 0.3 pg/ml, p<0.05). There was no difference in BP and deltaBV between the two groups during treatment. Plasma endothelin-1 was higher in hemodialysis patients and there was a continuous rise in plasma endothelin-1 during a session. Comparison of two groups of hemodialysis patients with and without s.c. rHuEPO-replacement treatment revealed a significantly higher plasma endothelin-1 concentration in patients with s.c. rHuEPO treatment. However, the elevated endothelin-1 levels were not accompanied by arterial hypertension.