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PubMed1983

[Determination of plasma hemoglobin in fresh plasma for therapeutic use by the TMB (3,3',5,5'-tetramethylbenzidine) method].

The maximum level of plasma hemoglobin in standard fresh frozen plasma before freezing has been mixed at 50 mg/l by french regulations. As no reference method is specified by the standards, we have adapted a technique where pseudo-peroxydasic activity of the hemoglobin is revealed by 3,3'-5,5', tetramethylbenzidine, a non carcinogenic chromogen derivate from benzidine. A 4 points reference scale is plotted for each plasma to be tested, thus reducing eventual interferences between pseudo-peroxydasic activity of the hemoglobin and other plasma proteins. We compared our modified technique to other existing ones: Vanzetti's extraction technique [8] whose main drawbacks are the use of flammable solvents and carcinogenic chromogen. Standefer's technique [10] involving plasma preincubation in H2O2 solution, which reveals protein interferences but is imprecise as it relies on only one reference point and does not show whether chromogen saturation is present. The level fixed by regulations is higher than the mean level indicated by our method, based on 118 samples of fresh plasma obtained by double centrifugation of whole blood collected on CPD as anticoagulant. It is also higher than the level indicated by the same method for plasma samples drawn from continuous or discontinuous flow cytapheresis (IBM 2997; Haemonetics V 50) using ACD - A or ACD - AA 16 as anticoagulant. Our method measures with precision low levels in plasma. Furthermore it is cheap, simple and easy to run in a blood transfusion center.

Benzidines↗
PubMed1988

Inhibition of gap junctional blockage by palmitoyl carnitine and TMB-8 in a rat liver epithelial cell line.

Exposure to 12-O-tetradecanoylphorbol-13-acetate (TPA) has been shown to inhibit gap junctional intercellular communication (GJIC) in many cell types in vitro. Using a scrape loading/dye transfer technique, TPA was shown to cause a dose-dependent and transient inhibition of GJIC in WB-F344, a normal rat liver epithelial cell line. Such a down-modulation of intercellular communication was found to be associated with an increase in protein kinase C (PKC) activity. Translocation of this activity to the particulate fraction occurred 10 min after exposure to 16 nM TPA and was consistent with the time course needed to inhibit GJIC. After 6 h exposure to TPA, essentially all the PKC activity was lost concurrent with the recovery of communication in these cells. During this time, the cells also became refractory to inhibition by further addition of TPA. Blockage of communication induced by TPA in WB cells was prevented by treating the cells with 23 microM palmitoyl carnitine for 1 h or 100 microM 8-N, N-(diethylamino)-octyl-3,4, 5-trimethoxybenzoate for 30 min. The results indicate that TPA transiently modulates GJIC in WB cells and PKC activation is possibly involved in blockage of communication in these cells.

Animals↗