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New biomedical devices with selective peptide recognition properties. Part 1: Characterization and cytotoxicity of molecularly imprinted polymers.

Rechichi A, Cristallini C, Vitale U, Ciardelli G, Barbani N, Vozzi G, Giusti P - J. Cell. Mol. Med. (2007 Nov-Dec)

Bottom Line: The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones).The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour.The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical Engineering, Politecnico di Torino, Torino, Italy.

ABSTRACT
Molecular imprinting is a technique for the synthesis of polymers capable to bind target molecules selectively. The imprinting of large proteins, such as cell adhesion proteins or cell receptors, opens the way to important and innovative biomedical applications. However, such molecules can incur into important conformational changes during the preparation of the imprinted polymer impairing the specificity of the recognition cavities. The "epitope approach" can overcome this limit by adopting, as template, a short peptide sequence representative of an accessible fragment of a larger protein. The resulting imprinted polymer can recognize both the template and the whole molecule thanks to the specific cavities for the epitope. In this work two molecularly imprinted polymer formulations (a macroporous monolith and nanospheres) were obtained using the protected peptide Z-Thr-Ala-Ala-OMe, as template, and Z-Thr-Ile-Leu-OMe, as analogue for the selectivity evaluation, methacrylic acid, as functional monomer, and trimethylolpropane trimethacrylate and pentaerythritol triacrylate (PETRA), as cross-linkers. Polymers were synthesized by precipitation polymerization and characterized by standard techniques. Polymerization and rebinding solutions were analyzed by high performance liquid chromatography. The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones). The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour. The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities. The cytotoxicity tests showed normal adhesion and proliferation of fibroblasts cultured in the medium that was put in contact with the imprinted polymers.

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(a) SEM of MIP 1+ resin showing the fused microgel particles of the macroporous monolith. (b) SEM of CP 1+ resin showing the fused microgel particles of the macroporous monolith. (c) SEM of MIP 1− resin showing aggregates of nanospheres. (d) SEM of CP 1− resin showing aggregates of nanospheres. (e) SEM of MIP 1− resin after suspension in methanol showing discrete and monodispersed nanospheres. (f) SEM of MIP 2− resin showing aggregates of nanospheres. (g) SEM of CP 2− resin showing aggregates of nanospheres. (h) SEM of MIP 2− resin after suspension in methanol showing small aggregates of nanospheres.
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fig01: (a) SEM of MIP 1+ resin showing the fused microgel particles of the macroporous monolith. (b) SEM of CP 1+ resin showing the fused microgel particles of the macroporous monolith. (c) SEM of MIP 1− resin showing aggregates of nanospheres. (d) SEM of CP 1− resin showing aggregates of nanospheres. (e) SEM of MIP 1− resin after suspension in methanol showing discrete and monodispersed nanospheres. (f) SEM of MIP 2− resin showing aggregates of nanospheres. (g) SEM of CP 2− resin showing aggregates of nanospheres. (h) SEM of MIP 2− resin after suspension in methanol showing small aggregates of nanospheres.

Mentions: SEM micrographs showed large fused aggregates of microgel particles interconnecting to form a labyrinth of macropores in the case of the more cross-linked resins as reported in Figure 1a. The images of MIP 1− showed a powder of nano-sized particles (Fig. 1c). Further investigation on samples obtained by suspension of the MIP 1− in methanol confirmed the spherical shape and the monodisperse size distribution of the nanospheres (Fig. 1e). Also for MIP 2− (Fig. 1f) a mass of particles was observed, but the particles seemed to form small aggregates, these were more clearly revealed by the analysis of the methanol suspension (Fig. 1h).


New biomedical devices with selective peptide recognition properties. Part 1: Characterization and cytotoxicity of molecularly imprinted polymers.

Rechichi A, Cristallini C, Vitale U, Ciardelli G, Barbani N, Vozzi G, Giusti P - J. Cell. Mol. Med. (2007 Nov-Dec)

(a) SEM of MIP 1+ resin showing the fused microgel particles of the macroporous monolith. (b) SEM of CP 1+ resin showing the fused microgel particles of the macroporous monolith. (c) SEM of MIP 1− resin showing aggregates of nanospheres. (d) SEM of CP 1− resin showing aggregates of nanospheres. (e) SEM of MIP 1− resin after suspension in methanol showing discrete and monodispersed nanospheres. (f) SEM of MIP 2− resin showing aggregates of nanospheres. (g) SEM of CP 2− resin showing aggregates of nanospheres. (h) SEM of MIP 2− resin after suspension in methanol showing small aggregates of nanospheres.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4401303&req=5

fig01: (a) SEM of MIP 1+ resin showing the fused microgel particles of the macroporous monolith. (b) SEM of CP 1+ resin showing the fused microgel particles of the macroporous monolith. (c) SEM of MIP 1− resin showing aggregates of nanospheres. (d) SEM of CP 1− resin showing aggregates of nanospheres. (e) SEM of MIP 1− resin after suspension in methanol showing discrete and monodispersed nanospheres. (f) SEM of MIP 2− resin showing aggregates of nanospheres. (g) SEM of CP 2− resin showing aggregates of nanospheres. (h) SEM of MIP 2− resin after suspension in methanol showing small aggregates of nanospheres.
Mentions: SEM micrographs showed large fused aggregates of microgel particles interconnecting to form a labyrinth of macropores in the case of the more cross-linked resins as reported in Figure 1a. The images of MIP 1− showed a powder of nano-sized particles (Fig. 1c). Further investigation on samples obtained by suspension of the MIP 1− in methanol confirmed the spherical shape and the monodisperse size distribution of the nanospheres (Fig. 1e). Also for MIP 2− (Fig. 1f) a mass of particles was observed, but the particles seemed to form small aggregates, these were more clearly revealed by the analysis of the methanol suspension (Fig. 1h).

Bottom Line: The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones).The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour.The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical Engineering, Politecnico di Torino, Torino, Italy.

ABSTRACT
Molecular imprinting is a technique for the synthesis of polymers capable to bind target molecules selectively. The imprinting of large proteins, such as cell adhesion proteins or cell receptors, opens the way to important and innovative biomedical applications. However, such molecules can incur into important conformational changes during the preparation of the imprinted polymer impairing the specificity of the recognition cavities. The "epitope approach" can overcome this limit by adopting, as template, a short peptide sequence representative of an accessible fragment of a larger protein. The resulting imprinted polymer can recognize both the template and the whole molecule thanks to the specific cavities for the epitope. In this work two molecularly imprinted polymer formulations (a macroporous monolith and nanospheres) were obtained using the protected peptide Z-Thr-Ala-Ala-OMe, as template, and Z-Thr-Ile-Leu-OMe, as analogue for the selectivity evaluation, methacrylic acid, as functional monomer, and trimethylolpropane trimethacrylate and pentaerythritol triacrylate (PETRA), as cross-linkers. Polymers were synthesized by precipitation polymerization and characterized by standard techniques. Polymerization and rebinding solutions were analyzed by high performance liquid chromatography. The highly cross-linked polymers retained about 70% of the total template amount, against (20% for the less cross-linked ones). The extracted template amount and the rebinding capacity decreased with the cross-linking degree, while the selectivity showed the opposite behaviour. The PETRA cross-linked polymers showed the best recognition (MIP 2-, alpha=1.71) and selectivity (MIP 2+, alpha'=5.58) capabilities. The cytotoxicity tests showed normal adhesion and proliferation of fibroblasts cultured in the medium that was put in contact with the imprinted polymers.

Show MeSH
Related in: MedlinePlus