Protein Complexation Onto Thermally Sensitive Microgels

Based on the effect of temperature on the protein adsorption, the oriented immobilization of proteins has been also investigated using N-(vinylbenzylimino)di-acetic acid (IDA) containing poly(N-isopropyl methacrylamide) microgel particles [32,33] as illustrated in Fig. 12. The aim of this work was to control the orientation and the direct immobilization of any proteic material bearing at least

FIG. 11 Protein desorbed amount (wt %) onto ammo-containing polystyrene core cross-linked poly(NIPAM) shell versus adsorption incubation time. Conditions: adsorption at pH 6.1 in the phosphate buffer (10 mM) and at 40°C (for 2 h), centrifugation and redispersion of the coated particles at 20°C in phosphate buffer (pH 6.1, 10 mM) at a given NaCl concentration (the incubation was performed over 17 h). (The hydrophilic thickness layer of DD4 latex is higher than that of DD11) [32].

FIG. 11 Protein desorbed amount (wt %) onto ammo-containing polystyrene core cross-linked poly(NIPAM) shell versus adsorption incubation time. Conditions: adsorption at pH 6.1 in the phosphate buffer (10 mM) and at 40°C (for 2 h), centrifugation and redispersion of the coated particles at 20°C in phosphate buffer (pH 6.1, 10 mM) at a given NaCl concentration (the incubation was performed over 17 h). (The hydrophilic thickness layer of DD4 latex is higher than that of DD11) [32].

six histidine tags. Then, by controlling the pH and salinity of the medium above the transition temperature, the adsorption efficiency enhances the affinity between the protein and the shrunken particles and, consequently, the complex-ation yield via additional divalent metals such as Zn2 +, Cu2 +, and Ni2 +. To favor the release of the specifically immobilized proteins onto chelating compound, the pH and the temperature are the key parameters of the immobilization process [32]. Such a process can be used in future immunoassays using modified antibodies (antibodies bearing histidine tag). For illustration, immobilization of the modified HIV-1 recombinant protein (RH24K) onto di-carboxylic-containing poly(NIPAM) microgel particles is given as a function of pH in Fig. 13.

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