Serum preparation

Serum sickness. This occurs when there is an excess of anhgen to antibody, resulting in the formation of soluble complexes. These may circulate and cause systemic reactions or be widely deposited in the kidneys, joints and skin. A rise in temperature, swollen lymph nodes, a generalized urticarial rash and painful swollen joints occur. The rcpeated administration of foreign serum (e.g. antidiphtheria serum or antitetanus serum prepared in horses) can lead to this condition due to antibodies being produced to the horse protein material. 

Materials required AChE preparations, commercial acetylcholinesterase, horse (blood) serum preparation, human (blood) serum preparation, or matrix biotests with these immobilized enzyme (see section 15.2.1)... 

True and pseudo-cholinesterase. The above serum preparations contained both the true and pseudo- cholinesterases of Mendel and Rudney.1 The effect of di-isopropyl phosphorofluoridate on these components was examined separately by means of the specific substrates described by Mendel, Mundel and Rudney,2 using the titration method described above. Phosphorofluoridate (5 x 10 8m) gave an inhibition of 57 per cent of the activity towards 00045m acetylcholine, 30 per cent of the activity towards 0-0005 m acetyl-/ methyl-choline, and 40 per cent of that towards 0-005 m benzoylcholine, after incubating the enzyme with the poison for 5 min. Thus in these experiments there appeared to be no appreciable difference in sensitivity of the true and pseudo-cholinesterases of horse serum to phosphorofluoridates. 

Polyclonal antibodies are produced by injecting an antigen into an animal in the presence of an adjuvant containing bacterial lipopolysaccharides that stimulate the immune system. Serum prepared from the blood contains several different classes of antibodies that interact with different domains in the antigen molecule, each of... 

Although the serum of animals immunized with small amounts of toxins has been used for many years to help people recover from diphtheria or to provide protection against snake or scorpion venom, there have been other situations where a specific antibody was needed in a concentrated form. Unfortunately, there were limits to what animal antibodies could do. First, the animal serum preparations, even with repeated injections, were mixtures of antibodies—not collections of many of the same antibody. That is just the way the immune system works in animals, including humans. In addition, there was always the danger of taking the serum from a diseased animal remember the case of Jim, the horse with tetanus ... 

Clarification by removal of casein with such agents as calcium chloride, acetic acid, cooper sulfate, or rennin has often been employed to obtain a serum more suitable for refractometric measurements. Obviously the composition, and hence the refractive index, of such sera will depend on the method of preparation. Furthermore, some of the serum proteins may be precipitated with the casein by some of the agents used, particularly if the milk has been heated. Refractive index measurements of such sera are not generally considered as satisfactory as freezing point measurements for detection of added water (David and MacDonald 1953 Munchberg and Narbutas 1937 Schuler 1938 Tell-mann 1933 Vleeschauwer and Waeyenberge 1941). Menefee and Overman (1939) reported a close relation between total solids in evaporated and condensed products and the refractive index of serum prepared therefrom by the copper sulfate method. Of course, a different proportionality constant would hold for each type of product. 

Serum Preparation Whole blood was collected after decapitation of the animal and allowed to clot at room temperature for 30 min. The serum was then separated from the clot by centrifugation at 5000 rpm at U C. Aliquots of the serum were immediately taken and refrigerated for the determination of glutamic-pyruvic transaminase and lactic dehydrogenase. An additional 0.5 ml of serum from each animal was treated with a few milligrams of sodium fluoride and refrigerated for later glucose determinations. The rest of the serum was frozen for the remainder of the clinical chemistry determinations. 

During the initial studies of hypro-protein, the levels were compared in plasma and serum prepared simultaneously from the same normal... 

Figure 7.1 Analysis of serum IgE concentrations following exposure of mice to allergens or vehicle. Groups of mice (n=6) were exposed to 50 pi of 25 per cent TMA or 1 per cent DNCB bilaterally on the shaved flanks. Control mice received identical treatment with vehicle (AOO) alone. Seven days later, 25 pi of the same test chemical at half the application concentration used previously, or an equal volume of vehicle alone, were applied to the dorsum of both ears. Fourteen days following the initiation of exposure, mice were exsanguinated by cardiac puncture and serum prepared. Serum IgE was measured using a sandwich ELISA. Results are expressed as mean serum IgE concentration in pg. ml 1 SE (where these exceeded 0.075 pg. ml-1). A summary of six independent experiments a-f.

In conclusion, the experimental protocol recommended presently is as follows. Groups of six BALB/c strain mice are exposed topically on both shaved flanks to 50 pi of one of three concentrations of the test material or to an equal volume of vehicle alone. Additional groups of mice receive 25 per cent TMA, 1 per cent DNCB or the same volume of 4 1 acetone olive oil, the vehicle of choice for these chemicals. Seven days later mice are treated on the dorsum of both ears with 25 pi of the same chemical at half the application concentration used previously, or with the same volume of vehicle alone. Fourteen days following the initiation of exposure, mice are exsanguinated by cardiac puncture and serum prepared and stored at -20°C until analysis (Hilton et al., 1995, 1996). This protocol is illustrated diagrammatically in Figure 7.2. 

The unknown may be simply a sodium chloride solution, or it may be serum. If serum is analyzed, then the standards should be prepared over a narrower concentration range to better bracket the unknown. Sodium in serum (approximately 140 meq/L, or 3200 ppm Na) may be determined by simple 1 100 dilution (e.g., 0.1 mL diluted to 10 mL) or 1 500 if required by the instrument. An alternative unknown is an artificial serum prepared by dissolving the following salts in water and diluting to 1 L ... 

Standard commercial control serum preparations (e.g., Versatol) may be used. Preparations with abnormal glucose concentrations are available, so the range of normal (ca. 90 mg/dL) to abnormal (300 mg/dL or greater) levels can be covered. 

Moeita H, Kita T Umeno M, Morita M, Yoshi-naga j and Okamoto K (1994) Analysis of serum elements and the contamination from devices used for serum preparation by inductivdy coupled plasma mass spectrometry. Sci Total Environ 151 9-17. 

Second-generation reference material refers to the serum preparation of Versieck et al. [49]. ID-MS, isotope dilution-mass spectrometiy NAA, neutron activation analysis FAAS, flame atomic absorption spectrometry ETAAS, electrothermal atomic absorption spectrometiy. 

G. Lovell and P.H. Corran, Determination of L-thyroxine in reference serum preparations as the o-phthalaldehyde-V-acetylcysteine derivative by reversed-phase liquid chromatography with electrochemical detection, J. Chromatogr., 1990, 525, 287-296. 

Booster injections with the same amount of peptide conjugate emulsified in Incomplete Freunds Adjuvant is given every second or third week. Blood can be collected following the second booster injection and serum prepared by standard procedures Normally, high-titer antipeptide antisera are obtained after two booster injections... 

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