Preparation of Antibodies

In the hybridoma method (F-ig. 6-14). a mouse or other small animal is sensiti/cd with an antigen. When a high enough titer of antibody against the selected antigen has been attained, the animal is sacrificed and its spleen cells are collected. The spleen cells contain a large number of B lymphocytes, and it is certain that some will he able to produce antigen-specific antibodies. Because the spleen cells are normal B lymph(x ytcs. they have a very short lifetime in cell cultures. Therefore, a method must be u.sed to extend their lifetime. 

Methods arc available for the devclopmcnl of MAbs with tf. ito lOO f human sequence. By using transgenic mice, all of the essential human antibody genes can be expre.s.scd. 

Gemtuzumab ozogamicin is indicated for the treatment of patients with CD33-positivc acute myeloid leukemia in frrst relapse among adults 60 years of age or older who are not considered candidates for cytotoxic chemotherapy. 

The indications for daclizumab arc prophylaxis of acute organ rejection in patients receiving renal tran.splants, as part of an immuno.suppressant regimen including cyclosporine and corticosteroids. The mechanism of action is the same as that of basiliximab. 

Murnmonab-CD3 blocks the function of T cells that are involved in acute renal rejection. Hence, it is indicated for the treatment of acute allograft rejection in heart and liver transplant recipients resistant to standard steroid thcrapie.s. 

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The preparation of antibodies specific for the individual plasma proteins has greatly facilitated their smdy, allowing the precipitation and isolation of pure proteins from the complex mixmre present in tissues or plasma. In addition, the use of isotopes has made possible the determination of their pathways of biosynthesis and of their turnover rates in plasma. 

Here, we describe the design and preparation of antibody supramolecular complexes and their application to a highly sensitive detection method. The complex formation between antibodies (IgG) and multivalent antigens is investigated. When an antibody solution is mixed with divalent antigen, a linear or cyclic supramolecule forms [26-29]. With trivalent antigens, the antibody forms network structures. These supramolecular formations are utilized for the ampH-fication of detection signals on the biosensor techniques. 

Preparation of Antibody Dendrimers and their Topoiogicai Structures... 

There are various techniques for the immobilization of proteins onto solid supports. To estimate their suitability for preparation of antibody sensing layers one have to consider some general requirements for the sensor function and application. 

Figure 6. Preparation of antibody multilayer using layer-by-layer deposition technique.

The following protocol represents a generalized method for protein thiolation using SATA. For comparison purposes, contrast the variation of this SATA modification method as outlined in Chapter 20, Section 1.1 for use in the preparation of antibody-enzyme conjugates. 

Blattler, W.A., Kuenzi, B.S., Lambert, J.M., and Senter, P.D. (1985b) New heterobifunctional protein cross-linking reagents and their use in the preparation of antibody-toxin conjugates. Photochem. Photobiol. 42, 231. 

Structure-Function Dissection and sequence analysis of phospholipases A2, 197, 201 cloning, expression, and purification of porcine pancreatic phospholipase A2 and mutants, 197, 214 preparation of antibodies to phospholipases A2, 197, 223 thermodynamics of phospholipase A2-ligand interactions, 197, 234 activation of phospholipase A2 on lipid bilayers, 197, 249 ... 

Use of Free Synthetic Peptides for the Preparation of Antibodies and T-Cells of Preselected Specificities... 

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