Phage MS2-based Signal-amplification Platform For ICPMS Bioanalysis

Together with element-tagging strategy,inductively coupled plasma mass spectrometry(ICPMS)has unique advantages in the quantitative analysis of biological materials.Introduction of exogenous metal tags,such as mercury(Hg),iodine(I),boron(B)and Gold(Au),especially lanthanides and their isotopes,provide a broader application perspective for ICPMS-based bioanalysis.Elemental complex tags used so far preferred simple and definitive stoichiometry,for example,1:1,toward targeted biomolecules,facilitating quantitative analysis of the targeted biomolecules via element-signal readout in ICPMS.When metallic nanoparticle was used,such as gold nanoparticle and quantum dot,the sensitivity of ICPMS-based bioanalysis could be very much enhanced due to the fact that one nanoparticle contains thousands of elemental atoms compared with those in one elemental complex tag.However,it brings some difficulties in such a way for absolute and accurate quantification of the biomolecules owing to the problem that the number of elemental atoms in nanoparticle could not be precisely controlled.Natural bio-nanoparticles like viral capsid proteins have regular size,good mono-dispersity and good biocompatibility,especially known number of easily modified amino acid residues,providing the possibility of labeling definite number of element-tags for signal-amplification,while selectivity of such a multiple element-tagged natural bio-nanoparticle toward targeted biomolecules can be realized through modification of a selective warhead.Therefore,we establish a natural nanoparticle-based platform for a more sensitive and selective bioanalysis using ICPMS.My thesis contains four chapters as below:In the first chapter,I introduced the state-of-art of element-tagging and signal-amplification strategies for ICPMS-based bioanalysis in recent years.In addition,the common natural bio-nanoparticles used in chemistry and life science are briefly summarized.In the second chapter,I used capsid protein of phage MS2 as bio-nanoparticles for signal-amplification of the ICPMS-based bioanalysis.I prepared and purified phage MS2 capsid protein,and modified azide groups onto the purified phage MS2 capsid protein for later copper-free clickable Eu-tagging.In the third chapter,I selected folic acid as a targeting molecule for fluorescence imaging of the folate receptors on tumor cells using fluorescent dye DBCO-Cy5 and folic acid modified MS2.In addition,MS2 was conjugated with DBCO-DOTA-Eu.Such Eu-tagged and folic acid modified MS2 was for preliminary study to count tumor cells.Finally,in the fourth chapter,I summarized what I did during my studies in master course.The prospect of more applications of the established signal-amplification strategy is also given.