An Improved Method for the Determination of the Plasma Volume with Evans Blue

Abstract

A new method for the spectrophotometric determination of Evans Blue in plasma is presented. The method is based on the precipitation of the non-albumin fraction of the plasma proteins with polyethylene glycol, in order to eliminate the variable background absorption of plasma at the wavelength of maximum absorbance of Evans Blue. The accuracy and precision of the method is excellent: at an Evans Blue concentration in plasma of 5 mg · 1 the coefficient of variation of the method is < 1 %. Five different procedures currently in use for the calculation of the plasma volume from the amount of indicator injected and the concentration at zero time (c 0 ) were compared. Extrapolation to zero time of the early part (10 to 60 min) of the log concentration vs time curve turned out to be the best method for the calculation of c 0 , and hence yields the best estimate of the plasma volume. Eine verbesserte Methode fur die Bestimmung des Plasmavolumens mit Evans BlueZusammenfassung: Eine neue Methode für die spektrophotometrische Bestimmung von Evans Blue wird beschrieben. Sie beruht auf der Fällung der Nicht-Albumin-Fraktion der Plasmaproteine mit Polyethylenglycol, um die variierende Untergrundabsorption von Plasma bei der Wellenlänge des Absorptionsmaximums von Evans Blue zu eliminieren. Richtigkeit und Genauigkeit der Methode sind hervorragend: bei einer Evans Blue-Konzentration von 5 mg/1 Plasma ist der Variationskoeffizient der Methode < 1%. Fünf verschiedene Verfahren, die gegenwärtig zur Ermittlung des Plasmavolumens aus der Menge an injiziertem Indikator und der Konzentration zum Zeitpunkt t = 0 (c 0 ) Verwendung finden, wurden verglichen. Extrapolation des frühen Teils (10-60 min) der log Konzentration gegen Zeit-Kurve auf die Zeit t = 0 erwies sich als die beste Methode für die Ermittlung von c 0 und ergibt deshalb den besten Wert für das Plasmavolumen.

Extended acclimatization is required to eliminate stress effects of periodic blood-sampling procedures on vasoactive hormones and blood volume in beagle dogs

“…On all occasions, animals were placed in slings, a catheter was inserted into the cephalic vein, and a 2 ml baseline blood sam ple was collected (in which HCT was measured). For actual PV determinati on, the cannula was¯ushed with saline and then a volume of Evan's blue dye (Sigma-Aldric h, Poole, UK) solution in saline (0.5 mg=ml), equivalent to one millilitre per kilogram body weight, was injected over a period of 30 s. Blood was collected (one millilitre) into lithium heparin at 10, 20, and 30 min post-infusion and PV was calculated from spectrophotometric measurements of Evan's blue dye in plasma treated with polyethylene glycol and using the baseline sam ple for blank and standard (Zweens & Frankena 1981). Procedures commenced at 12:30 h 15 min, and one and the sam e pair of technicians was used per dog (one to tend the animal and one to conduct the procedure).…”

Body Fluid Compartment Volumes in Chronically Hypoxemic Lambs

“…The exact amount of indicator injected was determined by weighing the syringe containing the indicator mixture before and after the injection. After treatment of the samples as described below, light absorbances were measured and converted to concentrations by using a calibration line that was determined before each set of measurements as described by Zweens et al (14)(15)(16). The Evans Blue concentration was measured in the samples withdrawn at 20, 30, 40, 50, and 60 min.…”

“…The concentration at time zero (coEB) was calculated by extrapolation from the monoexponential concentration versus time curve. Because Evans Blue is completely mixed with plasma 10 min after injection and subsequently disappears from plasma in a first-order fashion, the error made in cOEB by assuming monoexponential elimination between 20 and 60 min after injection is negligible (16). The plasma volume (V,) was calculated as:…”

Blood volume and body fluid compartments in lambs with aortopulmonary left-to-right shunts.

“…In brief, after induction of halothane anesthesia, we performed a thoracotomy through the fourth intercostal space and sutured a Goretex® conduit (6 mm (23). The indicators used were Evans blue, ferrocyanide, and deuterium oxide (D20), respectively (24)(25)(26). On the day of study the lamb was weighed, brought to the experimental room, and put in a sling.…”

“…Concentrations of Evans blue, sodium ferrocyanide, and D20 were measured spectrophotometrically as described by Zweens and coworkers (24,26,31 ). Evans blue concentration was determined in the samples that were withdrawn at 20, 30, 40, 50, and 60 min.…”