Antibodies to PAMAM dendrimers

Antibodies to PAMAM Dendrimers Reagents for Immune Detection, Patterning and Assembly of Dendrimers... 

Kunneman, D., Rowold, E. and Voliva, C. F. Recognition properties of antibodies to PAMAM dendrimers and their use in immune detection of dendrimers. Manuscript in preparation. 

S. C. Lee et al. Antibodies to PAMAM dendrimers reagents for immune detection, patterning and assembly of dendrimers, in Dendrimers and Other Derulritic Polymers. J.M.J. Frechet and D.A. TomaUa, Eds. John WUey Sons West Sussex, pp. 559-565, 2001. 

Monoclonal antibodies to sulfhydryl-terminated PAMAM dendrimers [9,10] could be bound to sulfhydryl-terminated PAMAM dendrimers under stoichiometric conditions favoring complexes of one antibody to one dendrimer. After binding, the exposed sulfhydryl surface may be derivatized to eliminate SH groups. Antibody and dendrimer complexes are then dissociated and the patterned dendrimers are isolated. Only areas masked by the antibody retain sulfhydryl groups, and can therefore be differentially derivatized (to a gold adduct, or to maleimide-linked macromolecules, for instance). 

Besides monoclonal antibodies, boronated PAMAM dendrimers can be targeted to tumor using epidermal growth factors [174,175], vascular endothelial growth factor [176], and folic acid, vitamin that is transported into cells via folate receptor mediated endocytosis [177]. 

Use of sulfo-NHS-LC-SPDP or other heterobifunctional crosslinkers to modify PAMAM dendrimers may be done along with the use of a secondary conjugation reaction to couple a detectable label or another protein to the dendrimer surface. Patri et al. (2004) used the SPDP activation method along with amine-reactive fluorescent labels (FITC or 6-carboxytetramethylrhodamine succinimidyl ester) to create an antibody conjugate, which also was detectable by fluorescent imaging. Thomas et al. (2004) used a similar procedure and the same crosslinker to thiolate dendrimers for conjugation with sulfo-SMCC-activated antibodies. In this case, the dendrimers were labeled with FITC at a level of 5 fluorescent molecules per G-5 PAMAM molecule. 

A common choice of crosslinker for this type of reaction is sulfo-SMCC, which has been used extensively for antibody conjugation (Chapter 20, Section 1.1). A better option for dendrimer conjugation is to use a similar crosslinker design, but one that contains a hydrophilic PEG spacer arm to promote dendrimer hydrophilicity after modification. Derivatization of an amine-dendrimer with a NHS-PEG-maleimide can create an intermediate that is coated with water-soluble PEG spacers. This modification helps to mask any potential for nonspecific interactions that the PAMAM surface may have, while providing terminal thiol-reactive maleimides for coupling ligands (Figure 7.10). 

Antibodies are powerful tools in pharmaceutical development, and as den-drimers are incorporated into various drugs and medical devices, antibodies such as described should be useful to study dendrimer biodistribution and pharmacokinetic properties. We have not attempted to detect PAMAM de-ndrimers in biological fluids (blood, urine, sputum, etc.), but expect that it would work. Additionally, the antibodies might be used in immunohistochemistry to study the tissue and cellular distribution of dendrimer therapeutics. 

Antibodies have been coupled1271 to starburst (PAMAM) dendrimers these reagents offer an attractive approach to the development of immunoassays. These dendritic reagents are stable and retain full immunological activity - both in solution as well as when bound to a solid substrate their analytical sensitivity is equal to or is better than that of the established methods. The dendritic approach to radial partition immunoassays has been shown to possess the best traits of both homogeneous and heterogeneous immunoassay formats. 

PAMAM dendrimer to target prostate-specific antigen (PSA) in prostate cancer. The antibody-conjugated dendrimer has been found to bind specifically to PSA positive cells (LNCaP human prostate cancer cell lines) but not with PSA negative cells (PC-3 human prostate cancer cell lines from bone) [50],... 

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