Measurement of Plasma Hemoglobin Peroxidase Activity

Abstract

We described a spectrophotometric method for measuring hemoglobin peroxidase activity in human plasma using o-dianisidine (o-DA) as the substrate and myeloperoxidase specific inhibitor 4-aminobensoic acid hydrazide (ruling out the probable contribution of myeloperoxidase to the measured parameter value). The optimal conditions (pH 5.5; 2 mM H2O2) have been determined, at which hemoglobin makes the main contribution to plasma oxidation of o-DA. A significant positive correlation between hemoglobin peroxidase activity measured by the spectrophotometric method and hemoglobin level measured by the pyridine hemochromogenic method has been detected (r=0.624; p<0.01) in plasma specimens from 16 donors. Plasma hemoglobin peroxidase activities were measured in healthy individuals and patients with type 2 diabetes mellitus and coronary heart disease. High plasma hemoglobin peroxidase activities in both groups of patients indicates disorders in the mechanisms of clearance of hemoglobin and its highly reactive derivatives and can serve as specific markers of diseases associated with oxidative stress.

Disposable immunoplatforms for the simultaneous determination of biomarkers for neurodegenerative disorders using poly(amidoamine) dendrimer/gold nanoparticle nanocomposite

“…Fig. 5 shows that significant differences were observed for the measured currents in the presence of Hb and HSA, similarly to that reported with other electrochemical immunosensors [28,68,69]. While Hb interference can be attributed to its well-documented endogenous peroxidase behavior [68], to our knowledge, the interference of HSA, although described in the literature [69], has not been explained.…”

“…5 shows that significant differences were observed for the measured currents in the presence of Hb and HSA, similarly to that reported with other electrochemical immunosensors [28,68,69]. While Hb interference can be attributed to its well-documented endogenous peroxidase behavior [68], to our knowledge, the interference of HSA, although described in the literature [69], has not been explained. Since our results show that the presence of HSA provoked an increase both in the specific and non-specific responses, we can hypothesize that HSA favors the possible non-specific adsorptions of the detection (TDP- or the secondary (HRP-anti-IgG) antibodies that occur in the absence of antigen.…”

High Presence of Extracellular Hemoglobin in the Periventricular White Matter Following Preterm Intraventricular Hemorrhage

“…In the current study, the inherent PO activity of Hb was utilized to identify and display the presence and distribution of both cellular and non-cellular retained Hb. The PO activity of Hb has previously mainly been utilized in plasma for qualitative or quantitative measurement of Hb (Grigorieva et al, 2013 ), however protocol for the use in histochemistry has been described (Strum and Karnovsky, 1970 ). The use of PO activity in tissue as an adjunct to specific Hb immunolabeling, which showed excellent co-distribution, for sensitive recognition of intracellular and extracellular Hb proved invaluable in the current study for achieving a spatial overview of the Hb distribution.…”

Easily Multiplexable Immunoplatform to Assist Heart Failure Diagnosis through Amperometric Determination of Galectin‐3

“…Regardless of the S/B ratio, the amperometric signals for Gal-3 were slightly higher when HSA, hemoglobin, Lp(a) and sAXL (bars 2, 3, 6 and 7, respectively) were present in the solution, thus indicating a sort of small interference from these non-target proteins. Moreover, in the presence of hemoglobin a slightly higher signal was obtained (Figure 4, white bars 3) likely due to its intrinsic peroxidase activity [44,45]. It is worth mentioning also that the supplier of the used immunoreagents for immunosensor development claimed no potential cross-reactivity or interference towards substances with similar molecular structures such as recombinant human Gal-1, Gal-2, Gal-4 Gal-7, Gal-8 and Gal-10 (all assayed at 50 ng mL À 1 ).…”