Hemolysis Histidin

Brydon and Roberts- added hemolyzed blood to unhemolyzed plasma, analyzed the specimens for a variety of constituents and then compared the values with those in the unhemolyzed plasma (B28). The following procedures were considered unaffected by hemolysis (up to 1 g/100 ml hemoglobin) urea (diacetyl monoxime) carbon dioxide content (phe-nolphthalein complex) iron binding capacity cholesterol (ferric chloride) creatinine (alkaline picrate) uric acid (phosphotungstate reduction) alkaline phosphatase (4-nitrophenyl phosphate) 5 -nucleotidase (adenosine monophosphate-nickel) and tartrate-labile acid phosphatase (phenyl phosphate). In Table 2 are shown those assays where increases were observed. The hemolysis used in these studies was equivalent to that produced by the breakdown of about 15 X 10 erythrocytes. In the bromocresol green albumin method it has been reported that for every 100 mg of hemoglobin/100 ml serum, the apparent albumin concentration is increased by 100 mg/100 ml (D12). Hemolysis releases some amino acids, such as histidine, into the plasma (Alb). 

Hence, noncrystallizing sugars (sucrose, trehalose) or polyols (dextran, polyethylene glycol) are added to the formulation to minimize cold denaturation during freeze-thaw or during lyophiliza-tion. Also, nonionic surfactants like polysorbates or Pluronics are included to prevent mAb adsorption at the interface of ice, air, or other product contact surfaces. Use of ionic surfactants like SDS is not advisable due to their potential to cause tissue irritation and hemolysis. Preformulation studies are important to identify the pH of maximum stability. For most mAbs, it is in the range of pH 5.0 to 6.5. Common buffers used to control the pH of the solution include phosphate, citrate, histidine, and glycine. 

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