Immunoglobulin solution

Fill the tubing to approx one-half of its capacity with the crude immunoglobulin solution from step 9 (see Note 13), and close the tubing with a knot or a dialysis tubing clip. 

Concentrate the immunoglobulin solution to approx 10 mg/mL under N2 on ice with gentle stirring. 

Determine the titer of immunoglobulin solution for the microorganism to be utilized in the assay. Select a dilution of the immunoglobulin that does not cause aggregation of the particles. If S. aureus or E. coli Bioparticles from Molecnlar Probes are nsed, follow the directions provided by the mannfacturer. 

Dilute alkaline phosphatase-conjugated polyvalent immunoglobulin solution 1 350 in washing buffer contmning 1% BSA. Add 200 pL/well of the substrate solution to all 60 inner wells that were treated with protamine sulfate. Incubate the microtiter plate at 37°C for 1 h. Remove the enzymewashing buffer as in Step 3. 

Place the immunoglobulin solution in Visking tubing and dialyze extensively against several changes of saline to remove sulfate ions. (Alternatively, remove sulfate by chromatography on a Se-phadex G-25 column.)... 

Check for residual sulfate ions by adding a few drops of the immunoglobulin solution to a tube containing a small volume of barium chloride solution. Any cloudiness indicates the presence of sulfate ions and the need for further dialysis. 

Measure the volume of immunoglobulin solution and calculate the protein concentration by measuring the absorbance of a 1 25 dilution at a wavelength of 280 nm using a cuvette of 1 cm path length. 

Measure the volume of the immunoglobulin solution and calculate the protein content as described above. 

Recentrifuge to remove any remaining cellulose, and decant the supernatant immunoglobulin solution. 

Place a measured volume of the immunoglobulin solution in a small beaker and cool to 4°. Place on magnetic stirrer. 

Add one-tenth volume of FITC solution dropwise while stirring the immunoglobulin solution at 4° (approximately 1 mg of dye per 100 mg of protein). 

Add 1 ml of immunoglobulin solution (previously dialyzed against saline) to the peroxidase solution. 

To prevent aseptic meningitis, it has been advised that intravenous immunoglobulin should be infused at a slow rate and that diluted immunoglobulin solutions should be used (58). Aseptic meningitis can be prevented by the administration of propranolol (41,58). In addition, prehydration and an antihistamine have been helpful in some patients (41,58). 

Another exanple of such non-conventional, innovative methods is applying immunoglobulin solutions films on the surface of carbon steel and stainless steel that has been shown to prevent the adherence of Pseudomonas jluorescens on these metallic surfaces, thus inhibiting biofilm formation [4]. However, this method would not become popular in industry, perhaps because of reasons such as... 

The development of freeze-drying for the production of blood derivatives was pioneered during World War II (96,97). It is used for the stabilization of coagulation factor (98,99) and intravenous immunoglobulin (IgG iv) products, and also for the removal of ethanol from intramuscular immunoglobulin (IgG im) solutions prior to their final formulation (Fig. 2). 

The resuspended and formulated Fraction II precipitate normally contains some aggregated IgG and trace substances that can cause hypotensive reactions in patients, such as the enzyme prekail ikrein activator (186). These features restrict this type of product to intramuscular adininistration. Further processing is required if products suitable for intravenous adininistration are required. Processes used for this purpose include treatment at pH 4 with the enzyme pepsin [9001-75-6] being added if necessary (131,184), or further purification by ion-exchange chromatography (44). These and other methods have been fiiUy reviewed (45,185,187,188). Intravenous immunoglobulin products are usually suppHed in the freeze-dried state but a product stable in the solution state is also available (189). 

Monkos K, Turczynski B. 1999. A comparative study on viscosity of human, bovine and pig IgC immunoglobulins in aqueous solutions. International Journal of... 

An electrode in which an antibody or an antigen/hapten is incorporated in the sensing element is termed an immunoelectrode . The potential response of the immuno-electrode is based on an immunochemical reaction between the sensing element of the electrode and antibody or antigen/hapten in the sample solution. One example of such an electrode is the polymer membrane electrode shown in Fig. 7. The selective response of this electrode to specific immunoglobulins is based on the interaction between antibody in solution and an antigen-ionophore complex in the membrane ... 

Secondary antibody and determination. A secondary antibody labeled with an enzyme is added which binds to the primary antibody that is bound to the coating antigen. If the primary antibody were produced in a rabbit, an appropriate secondary antibody would be goat anti-rabbit immunoglobulin G (IgG) conjugated with horseradish peroxidase (HRP) (or another enzyme label). Excess secondary antibody is washed away. An appropriate substrate solution is added that will produce a colored or fluorescent product after enzymatic conversion. The amount of enzyme product formed is directly proportional to the amount of first antibody bound to the coating antigen on the plate and is inversely proportional to the amount of analyte in the standards. 

Cellulose can be activated by CDI and coupled with the amino groups of peptides or immunoglobulins in aqueous alkaline solution to give immobilized peptides or antibodies such as the immunoglobulin IgG [210] (see also Section 6.2) ... 

Figures 9 and 10 represent a selected comparison of amide V and I+II FTIR and VCD for four proteins in D2O solution. Of these, myoglobin (MYO) has a very high fraction of a-helix, immunoglobulin (IMU) has substantial /1-sheet component, lactoferrin (LAF) has both a and j3 contributions, and a-casein (CAS) supposedly has no extended structure. The FTIR spectra of these proteins change little, the primary difference...

With mixing, add a quantity of the sulfo-NHS-biotin solution to the protein solution to obtain a 12- to 20-fold molar excess of biotinylation reagent over the quantity of protein present. For instance, for an immunoglobulin (MW 150,000) at a concentration of 10 mg/ml, 20 pi of a sulfo-NHS-biotin solution (containing 8 X 10-4 mmol) should be added per ml of antibody solution to obtain a 12-fold molar excess. For more dilute protein solutions (i.e., 1-2 mg/ml), increased amounts of biotinylation reagent may be required (i.e., 20-fold molar excess or more) to obtain similar incorporation yields as when using more concentrated protein solutions. 

Add 0.3 mg of sulfo-NHS-SS-biotin (Thermo Fisher) to each ml of the antibody solution. To measure out small amounts of the biotinylation reagent, it may be first dissolved in water at a concentration of at least 1 mg/ml. Immediately transfer the appropriate amount to the antibody solution. This level of sulfo-NHS-SS-biotin addition represents about an 8-fold molar excess over the amount of antibody present. This should result in a molar incorporation of approximately 2-4 biotins per immunoglobulin molecule. 

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