Pretreatment of Serum Containing Hemoglobin Vesicles (Oxygen Carriers) to Prevent Their Interference in Laboratory Tests

Abstract

Hemoglobin vesicles (HbV, diameter: 251 +/- 81 nm) are artificial oxygen (O2) carriers encapsulating concentrated hemoglobin (Hb) solution with phospholipid bilayer membrane, and their O2 transporting ability in vivo has been extensively studied. It is important to clarify the interference of the HbV suspension in clinical laboratory tests performed on serum and to establish a pretreatment method to avoid such an interference. The HbV suspension, acellular Hb solution ([Hb] = 10 g/dl) or saline, was mixed with a pooled human serum at various ratios up to 50 vol% ([Hb] = 5 g/dl), and the magnitude of the interference effect of HbV and Hb on 30 analytes was studied. The mixture of the HbV suspension and serum was ultracentrifuged (50,000 g, 20 min) to remove the HbV particles as precipitate, and the supernatant was analyzed and compared with the saline control group. The HbV particles were also removed by centrifugation (2,700 g, 30 min) in the presence of dextran (Mw 200 kDa). The HbV suspension showed considerable interference effects in most analytes. The majority of these effects was more serious than those of the acellular Hb solution. These findings are thought to be due to the light absorption of Hb in HbV and/or the light scattering generated in the suspension that interferes with the colorimetric and turbidimetric measurements. The components of HbV may also interfere with the chemical reactions of the studied assays. However, removal of the HbV from the supernatant diminished the interference in most of the assays: this is an advantage of HbV in comparison with acellular chemically modified Hb solutions.

Repeated Injection of High Doses of Hemoglobin-Encapsulated Liposomes (Hemoglobin Vesicles) Induces Accelerated Blood Clearance in a Hemorrhagic Shock Rat Model

“…Every day after injection, blood was collected from the tail vein under ether anesthesia. Plasma was collected after centrifugation (3000g, 5 min), and the supernatant was subsequently ultracentrifuged to remove intact HbV (50,000g, 30 min) (Sakai et al, 2003). The supernatant was collected as the plasma sample and was stored at Ϫ80°C until used.…”

Hemoglobin encapsulation in vesicles retards NO and CO binding and O2release when perfused through narrow gas-permeable tubes

“…After a steady flow was attained, three measurements were performed. Because of the light scatter by fine particles, the absorbance of the HbVs at a shorter wavelength was slightly higher than that at a longer wavelength (41). In the spectra of the 100% deoxy-Hb and 100% oxygenated Hb-containing samples, two isosbestic points (522 and 586 nm) in the Q band of Hb were connected by a straight line as the baseline (Fig.…”

Evaluation of Safety and Efficacy of Hemoglobin-Vesicles and Albumin-Hemes

“…However, HbV can be simply removed from blood plasma either by ultracentrifugation or centrifugation in the presence of a high-molecular-weight dextran to enhance precipitation. We can obtain a very clear supernatant for accurate analyses [35]. This is one advantage of HbV in comparison with acellular Hb solutions.…”

“…We further examined the safety profile of HbV such as cardiovascular responses, pharmacokinetics, influence on RES, influence on clinical measurements and daily repeated infusion [29][30][31][32][33][34][35][36][37].…”