A Virus-like Element-tagged Nanoparticle ICPMS Signal Multiplier

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A Virus-like Element-tagged Nanoparticle ICPMS Signal Multiplier: Membrane Biomarker Mediated Cell Counting Rong Yuan, Fuchun Ge, Yong Liang, Yang Zhou, Limin Yang, and Qiuquan Wang Anal. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.analchem.9b00749 • Publication Date (Web): 02 Apr 2019 Downloaded from http://pubs.acs.org on April 2, 2019

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Analytical Chemistry

A Virus-like Element-tagged Nanoparticle ICPMS Signal Multiplier: Membrane Biomarker Mediated Cell Counting Rong Yuan,† Fuchun Ge,† Yong Liang,† Yang Zhou,† Limin Yang,† and Qiuquan Wang*,†,‡ † Department of Chemistry & the MOE Key Laboratory of Spectrochemical Analysis and Instrumentation, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005, China ‡ State Key Lab of Marine Environmental Science, Xiamen University, Xiamen 361005, China E-mail: [email protected] ABSTRACT: Although rare cancerous cells are considered as more objective indications for a precise early diagnosis of cancers, accurate counting of them still is a spirited challenging. We reported a signal multiplication strategy by constructing element-tagged virus-like nanoparticles (VLNPs) with precise number of atoms for a membrane biomarker mediated higher sensitive cell counting using inductively coupled plasma mass spectrometry (ICPMS). Typical bacteriophage MS2 was exemplified to demonstrate the effectiveness of the element-tagged VLNPs as signal multipliers. Dibenzylcyclooctynepoly(ethylene glycol)-folate (DBCO-PEG-FA) and DOTA-Eu complex tag modified (FA-PEG)69-MS2-(DOTA-Eu)965 targeted the folate receptor (FR) on KB cells, as low as sub-zmol FRs could be quantified by 153Eu-species unspecific isotope dilution ICPMS, allowing us to be able to count at least 5 KB cells. While more than 2197 KB cells were needed to give a significant ICPMS signal using FA-PEG-DOTA-Eu, demonstrating more than two orders of magnitude signal multiplication and resulting in totally 4.0  108 times signal amplification relative to one KB cell. We believe that such a signal multiplication strategy can be expanded to quantify and count other membrane biomarkers and their host cells using various VLNPs modified with different kinds and precise numbers of elements and guiding groups. In this way, prescribed multiples of signal amplification can be realized for a more accurate ICPMSbased quantitative bioanalysis because targeted molecules/cells in a complicated biological system might exist in orders of magnitude wide concentration range.

Sensitivity and selectivity are the most important premises to accomplish an accurately quantitative bioanalysis. With the advantages of at least pg mL-1 level limit of detection and 0.5 atomic mass unit resolution as well as free of sample matrix, hard ionization inductively coupled plasma mass spectrometry (ICPMS) has evolved into a promising bioanalytical tool since the beginning of this century,1-3 thanks to the continuous developments of chemoselective and biospecific elementtagging strategies.4-11 Compared with the most popular soft ionization ESI/MALDI-based mass spectrometry, multiple proteins and nucleic acids could be quantified with only one isotopic element standard using ICPMS via selective labeling of element-complex tags.12-17 Cells could also be counted when their specific membrane molecules were labeled with the element-complex tags, resulting in the membrane molecule quantity times signal amplification relative to one cell itself.1821 Biomolecule analysis and cell counting sensitivity could be further improved when element-loaded engineered nanoparticles and polymers were used.22-31 They did well for a relative signal amplification, however, were particularly prone to some uncertainty for an absolutely quantitative bioanalysis because the number of elemental atoms and/or ions in the nanoparticles and polymers was somewhat difficult to control within a tolerant range.32,33 For example, the most widely used Au nanoparticles prepared with conventional methods using

citrate reduction of HAuCl4 spanned 10% variation from small to large particles; and the lanthanide-containing polymer particles about 100 nm diameter coped with the size distribution range of 0.03 to 0.209 polydispersity index, the number of the lanthanide ions chelated by the individual particles was not easy to accurately control. These uncertainties are actually more serious when rare tumor cells need to be counted at the early stage of cancers. Although smaller sized nanoclusters such as Au24Peptide8 had definite number of Au,34 the multiple of signal amplification was obviously limited to tens of times. In the latest decade, natural virus-like nanoparticles (VLNPs) with precisely viral structural protein capsids have been used as vaccine carriers, therapeutic and imaging agents as well,35,36 as their capsid proteins not only have numerous known chemically modifiable amino acid residues but also are highly selforganized. We hypothesize that the surface of VLNPs can be precisely labeled with definite number of the element-complex tags besides targeting groups, and the element-tagged VLNPs may serve as signal multipliers for a more sensitive thus accurate ICPMS-based bioanalysis. However, the feasibility of such element-tagged VLNPs to be used as potential ICPMS signal multipliers has yet remained to be explored. Typical bacteriophage MS2 was thus chosen as an example of VLNPs in this study to demonstrate the possibility of

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element-tagged VLNPs signal multiplication due to its biosafety, easy production in laboratory, monodisperse property and especially tolerance toward extensive chemical modifications. MS2 capsid is comprised of 180 protein monomers of 13728 Da molecular weight each, and has a sphere-shaped conformation with the outer and inner diameters of 27 and 21 nm. Moreover, there are 32 pores of 18 Å in the capsid shell,37 providing accesses to the interior surface modification. Each protein monomer possesses seven reactive amino groups including three lysine and one terminal amino groups located on the exterior surface and three other lysine amino groups on the interior surface (https://www.rcsb.org/structure/2MS2). Bearing these in mind, we first prepared empty MS2 capsid by removing its RNA genome (Figure S1 in Supporting Information), exposing both the exterior and interior surface for NHS-PEGn-N3 ester modification via the typical acylation reaction between NHS and amino group (-NH2). The element-complex tag, DBCODOTA-Eu (Figure S2a), could be conjugated onto the surface of the obtained azido-MS2 capsid via the strain promoted alkyne-azide cycloaddition (SPAAC) click reaction between DBCO and -N3 to obtain MS2-DOTA-Eu. On the other hand, folate receptor (FR), a glycosylphosphatidylinositol membraneanchored protein, on FR++-positive human nasopharyngeal carcinoma KB cells was exemplified to show the effectiveness of MS2-DOTA-Eu for ICPMS-based cell membrane molecule mediated cell counting. In order to make MS2-DOTA-Eu has the targeting ability toward KB cells through the specific interaction between the FR on KB cells and folic acid (FA), FAcontaining DBCO-PEG-FA was modified on MS2-DOTA-Eu to generate FR-targeted FA-PEG-MS2-DOTA-Eu ICPMS signal multiplier (Figure 1 and Scheme S1). O NH2

O

O N

𝒎

N3 O n On = 1, 4, 8, 12

H N

O O

𝟓𝟎 𝐦𝐌 𝐏𝐁𝐒, 𝐩𝐇 = 𝟕. 𝟖

𝑴𝑺𝟐

n N3

𝒎

𝑫𝑩𝑪𝑶 − 𝑷𝑬𝑮 − 𝑭𝑨 𝑫𝑩𝑪𝑶 − 𝑫𝑶𝑻𝑨 − 𝑬𝒖 𝟓𝟎 𝐦𝐌 𝐇𝐄𝐏𝐄𝐒, 𝐩𝐇 = 𝟔. 𝟖

𝑴𝑺𝟐 − 𝑵𝟑

H N

O O

N O

H H N PEG N O

O

N H

N H

𝑫𝑩𝑪𝑶 − 𝑷𝑬𝑮 − 𝑭𝑨

N N

OH

N

NH

O

O

N N

NH2

N O

O N

O

Eu N

O

N

O N

PEG-FA

𝒎𝟏 NH O

COOH O

nN N N

O

n N N N

might randomly enter the interior as existing stochastically in a linear configuration. Moreover, it should be noted that although the outer surface -NH2 groups could be easily modified by NHS-PEGn-N3, the interior K43, K57 and K61 that configure adjacently on the internal surface (https://www.rcsb.org/structure/2MS2) exhibited somewhat steric hindrance to prevent from being completely modified. Fortunately, the modification reproducibility was acceptable with less than RSD of 3.8% (n = 11). Owing to 1034  40 DBCO-DOTA-Eu was modified on one N3-MS2 capsid (Figure 2b, c and e), the multiple of signal amplification was more than 3 orders of magnitude larger compared with just using DBCO-DOTA-Eu complex tag (Figure 2a). This was determined by 153Eu-species unspecific isotope dilution (SUID) ICPMS, which was coupled with size-exclusion chromatography (SEC) to get rid of untagged free DBCODOTA-Eu (Table S1). Clearly, such a strategy can be applied not only to count viruses themselves, but also can be expected to quantify cell membrane protein and count its host cells when MS2-DOTA-Eu links with a cell membrane protein-targeting group.

O N

DOTA-Eu

𝒎𝟐 (𝑭𝑨 − 𝑷𝑬𝑮) 𝒎𝟏 −𝑴𝑺𝟐 − (𝑫𝑶𝑻𝑨 − 𝑬𝒖) 𝒎𝟐

O O

𝑫𝑩𝑪𝑶 − 𝑫𝑶𝑻𝑨 − 𝑬𝒖

Figure 1. Construction of Eu-tagged MS2 with folic acid targeting group. The amino group (-NH2) on MS2 was first reacted with NHS-PEGn-N3 to prepare MS2-N3 (n = 1, 4, 8 and 12, respectively). And then DBCO-PEG-FA and DBCO-DOTA-Eu were simultaneously modified to prepare FA-PEG-MS2-DOTA-Eu.

An ideal ICPMS signal multiplier should contain more reporting atoms/ions. The number of Eu decorated in MS2 capsid depends mostly on the efficiency of NHS-PEGn-N3 modification because the following SPAAC38 click conjugation with excess DBCO-DOTA-Eu (16 Å) almost proceeds completely. At the beginning, we thought that the longer PEG length in NHS-PEGn-N3, the more flexible to manipulate the surface of MS2 capsid owing to the better solubility and easier to be tagged. However, when using different length NHS-PEGnN3 (n = 1, 4, 8, 12) to modify MS2 capsid, we found that NHSPEG1-N3 (15 Å) was the best compared with NHS-PEG4-N3 (25 Å), NHS-PEG8-N3 (40 Å) and NHS-PEG12-N3 (54 Å). MALDITOF-MS results (Figure S3) indicated that six of the seven amino groups on one monomer of MS2 capsid could be modified by NHS-PEG1-N3, while no more than five by the other NHS-PEGn-N3 (n = 4, 8 and 12). These are ascribed to the 18 Å pore-size in the capsid shell that restricts the bigger molecules to come into the interior, even though some of them

Figure 2. 153Eu-SUID-SEC/ICPMS quantification and H3[P(W3O10)4] negatively stained TEM imaging of MS2-(DOTAEu)1034. Eu signal intensities of MS2-(DOTA-Eu)1034 and DBCODOTA-Eu complex (10 nM each) (a); signal intensities of 151Eu (black) and 153Eu (red) (b) and their mass flow chromatogram (c). TEM imaging (scale bar 50 nm) of MS2-N3 (d), MS2-(DOTAEu)1034 (e), and (FA-PEG)69-MS2-(DOTA-Eu)965 (f).

Subsequently, we modified MS2-N3 (Figure 2d) using DBCO-PEG-Eu together with DBCO-DOTA-FA in order to demonstrate the signal multiplication ability of MS2-DOTA-Eu for quantifying FRs and counting KB cells. A 15 ethylene glycol unit-containing PEG was used in DBCO-PEG-FA molecule to 1) have a longer arm favorable for targeting FR thus catching the cells and 2) merely label the exterior surface of MS2-N3 as well as 3) avoid the nonspecific adsorption. Considering that both of DBCO-PEG-FA and DBCO-DOTAEu have the same clickable DBCO group, we could control the number of DBCO-PEG-FA and/or DBCO-DOTA-Eu modified on MS2-N3 by adjusting their actually used concentration ratio. The number of DBCO-DOTA-Eu decreased from 1034  40 to 442 ± 14 (n = 7) along with the increase in the ratio of DBCOPEG-FA to DBCO-DOTA-Eu from 0 to 1/2, while that of DBCO-PEG-FA increased from 0 to 592  19 determined using 153Eu-SUID-SEC/ICPMS (Table S1). To obtain more Eu on FA-PEG-MS2-DOTA-Eu while maintain the targeting ability to FRs on KB cells, we chose 1/24 of DBCO-PEG-FA to DBCO-DOTA-Eu. In this case, FA-PEG-MS2-DOTA-Eu had (965  13) Eu and (69  1) FA (n = 7). Such a (FA-PEG)69MS2-(DOTA-Eu)965 (Figure S4) still posed the sphere


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Analytical Chemistry conformation (Figure 2f), and its -potential (-24.8 ± 0.5 mV) was almost the same as that of the native MS2 capsid (-24.3 ± 1.1 mV) (Table S2). Moreover, its biocompatibility was so good that it did not influence the cell viability (Figure S5). Before evaluating its targeting ability and signal multiplication capability, we must answer a question: how many FA on each (FA-PEG)69-MS2-(DOTA-Eu)965 simultaneously interacted with FRs on the cell surface. We thus synthesized FA-PEGDOTA-Eu complex tag (Figure S2c), in which the PEG used has the same length as that in (FA-PEG)69-MS2-(DOTA-Eu)965. We used FA-PEG-DOTA-Eu as a FR-targeted signal readout tag to determine the amount of FRs expressed on KB cells using ICPMS. Based on the fact that FA can specifically interact with FR with 1:1 stoichiometry,39 the obtained results indicated that average amount of the FR on one KB cell was 1.38 × 10-18 mol (RSD = 2.1%, n = 5) corresponding to 8.30 × 105 FR molecules, during which 5.0  106 KB cells measured using a blood counting chamber were used. This means that nearly six orders of magnitude signal amplification relative to one KB cell itself could be achieved when we determined the Eu tagged to FRs for counting its host KB cells. The targeting ability and signal multiplication capability of (FA-PEG)69-MS2-(DOTA-Eu)965 were then investigated. The specific targeting ability toward FRs on KB cells was first verified by the labeling and blocking experiments using 500 M free FA molecules. Significant Eu signal was determined from the tagged KB cells while negligible Eu signal from the FA-blocked cells (Figure S6a), suggesting that the FA on (FA-PEG)69-MS2-(DOTA-Eu)965 could specifically target FRs on the cells. 2.0  105 KB cells were incubated with different amounts of (FA-PEG)69-MS2(DOTA-Eu)965 from 2 to 100 nM and different time from 15 min to 4.0 h under 4 C rather than 37 C for avoiding any cell endocytosis. The obtained results (Figure S7) indicated that the FRs on KB cell membrane could be completely labeled within 2.5 h using 50 nM (FA-PEG)69-MS2-(DOTA-Eu)965. The average amount of Eu tagged on one KB cell was determined to be 6.57 × 10-16 mol (RSD = 2.0 %, n = 5). If only one FA on each (FA-PEG)69-MS2-(DOTA-Eu)965 interacted with one FR on KB cell surface, it would be 6.8 × 10-19 (6.57 × 10-16/965) mol FRs on each KB cell surface. However, compared to 1.38 × 10-18 mol FRs/cell obtained using FA-PEG-DOTA-Eu, there was close to two times decrease of FRs. These two times decrease implied that 2 out of 69 FA on each (FA-PEG)69-MS2(DOTA-Eu)965 interacted with 2 FRs on one KB cell. This might be ascribed to the size of (FA-PEG)69-MS2-(DOTA-Eu)965 sphere is the much smaller compared with the bigger FRoverexpressed KB cell (15-20 m). The average amount of FR on each KB cell was thus determined as 1.36 × 10-18 [2 × (6.57 × 10-16/965)] mol (RSD = 2.0 %, n = 5), being well in agreement with that determined using FA-PEG-DOTA-Eu. More importantly, when labeling the FRs on KB cells with (FAPEG)69-MS2-(DOTA-Eu)965 for counting the cells, we gained 482.5 (965/2) times more signal multiplication. Totally 4.0  108 (8.30  105  482.5) times signal amplification relative to one KB cell itself was achieved, demonstrating the signal multiplication capability of (FA-PEG)69-MS2-(DOTA-Eu)965. In this way, we were able to directly count at least 5 KB cells per mL for the first time (Figure 3a) according the detection limit (3) of Eu (0.477 pg mL-1) using ICPMS under a conventional bulk sample introduction manner. While more than 2197 KB cells were needed to give a significant ICPMS signal using FA-PEG-DOTA-Eu (Figure 3b). This signal multiplication capability was also confirmed using less FR-

expressed, FR+-positive Hela cells (Figure S6a), showing that the amount of FRs on Hela cells was 2.13 × 10-19 mol/cell (RSD = 1.2%, n = 5), which was in agreement with the value reported before, one order of magnitude lower than that on KB cell.40 Even though, at least 34 Hela cells could be theoretically counted considering the FA amount expressed and the (FAPEG)69-MS2-(DOTA-Eu)965 signal multiplication outcome. It is worthy of noting that although A549 was traditionally considered as a FR--negative alveolar adenocarcinoma cell, FR on A549 cells was still determined as 4.44 × 10-20 mol FR per cell (RSD = 5.0 %, n = 5) and thus at least 163 A549 cells could be counted. Such a small amounts of FR were hardly detected by fluorescent imaging under LCSM with the FA-PED-MS2Cy5, in which the popular red color DBCO-Cy5 reporting molecule instead of DBCO-DOTA-Eu was modified on the MS2 capsid (Scheme S2 and Figure S6).

Figure 3. Relationship of 153Eu intensity (153Euspike  99.9% + 153Eu 153Eu-SUID-ICPMS Sampple  52.19%) determined using against the number of KB cells labeled with (FA-PEG)69-MS2(DOTA-Eu)965 (blue) and FA-PEG-DOTA-Eu (red) (n = 5).

Conclusively, we fabricated a Eu-tagged MS2 signal multiplier with precise number of Eu to improve the sensitivity of ICPMS-based bioanalysis for the first time. When FRtargeted FA was modified on the signal multiplier, as low as sub-zmol level FR could be quantified, thus several to hundreds of FR-expressed cancer cells could be counted without preconcentration procedures using ICPMS under the conventional bulk sample introduction manner. Such a signal multiplication strategy using element-tagged VLNPs can be very much expected to apply to simultaneous counting and profiling of other cells using mass cytometry,41-43 when various VLNPs are chemically tagged with diverse elements/isotopes and modified with various targeting groups. Furthermore, the VLNPs’ scaffold can be manipulated in biological ways. Uncanonical amino acids with biological specific targeting and element-modifiable groups can be genetically assembled on the capsids of VLNPs.44,45 Such element-tagged VLNPs will display prescribed multiples of signal amplification for the targeted molecules/cells that might exist in orders of magnitude wide concentration range in a complex biosystem, to accomplish the goal of simultaneous quantification of biological meaningful molecules on single cells for cell profiling and counting, which has been dreamed about but never been realized before.

ASSOCIATED CONTENT Supporting Information The Supporting Information is available free of charge on the ACS Publications website at DOI: Materials and instrumentation, synthesis and characterization of DBCO-DOTA-Eu and FA-PEG-DOTA-Eu, MS2, MS2-N3, MS2(DOTA-Eu)1034, (FA-PEG)69-MS2-(DOTA-Eu)965 and FA-PEF-



MS2-Cy5, FA-FR interaction based cell labeling and counting (Scheme S1-2, Figure S1-7 and Table S1-4) (PDF).

AUTHOR INFORMATION Corresponding Author * E-mail: [email protected]; Fax: +86 (0)592 2187400.

ORCID Qiuquan Wang: 0000-0002-5166-4048

Notes The authors declare no competing financial interest.

ACKNOWLEDGMENT This work was financially supported by the National Natural Science Foundation of China (Grants 21535007, 21874112 and 201475108), the National Basic Research 973 Program (Grant 2014CB932004), and the National Science and Technology Basic Work (2015FY111400). We thank the Foundation for Innovative Research Groups of the National Natural Science Foundation of China (Grant 21521004) and Program for Changjiang Scholars and Innovative Research Team in University (PCSIRT, Grant IRT13036) for partly financial support. We thank Prof. Gang Liu (Center for molecular imaging and translational medicine, school of public health, Xiamen University) and his students Pengfei Zhang, En Ren, Yunming Zhang for their generous assistance during bacteriophage MS2 preparation process and helpful discussions.

REFERENCES (1) Zhang, C.; Wu, F. B.; Zhang, Y. Y.; Wang, X.; Zhang, X. R. J. Anal. At. Spectrom. 2001, 16, 1393-1396 (2) Zhang, C.; Zhang, Z. Y.; Yu, B. B.; Shi, J. J.; Zhang, X. R. Anal. Chem. 2002, 74: 96-99. (3) Baranov, V. I.; Quinn, Z.; Bandura, D. R.; Tanner, S. D. Anal. Chem. 2002, 74: 1629-1636. (4) Bettmer, J.; Montes-Bayón, M; Ruiz Encinar, J.; Sanchez, M. L.; Fernández de la Campa, M. D. F.; Sanz-Medel, A. J. Proteomics 2009, 72, 989-1005. (5) Tholey, A.; Schaumlöffel, D. TrAC, Trends Anal. Chem. 2010, 29, 399-408. (6) Sanz-Medel, A.; Montes-Bayón, M.; Bettmer, J.; FernándezSanchez, M. L.; Ruiz Encinar, J. TrAC, Trends Anal. Chem. 2012, 40, 52-63. (7) Yan, X. W.; Yang, L. M.; Wang, Q. Q. Anal. Bioanal. Chem. 2013, 405, 5663-5670. (8) Schwarz, G.; Mueller, L.; Beck, S.; Linscheid, M. W. J. Anal. At. Spectrom. 2014, 29, 221-233. (9) de Bang, T. C.; Husted, S. TrAC, Trends Anal. Chem. 2015, 72, 45-52. (10) Liang, Y.; Yang, L. M.; Wang, Q. Q. Appl. Spectrosc. Rev. 2016, 51, 117-128. (11) Liu, R.; Zhang, S. X.; Wei, C.; Xing, Z.; Zhang, S. C.; Zhang, X. R. Acc. Chem. Res. 2016, 49, 775-783. (12) Yan, X. W.; Xu, M.; Yang, L. M.; Wang, Q. Q. Anal. Chem. 2010, 82, 1261-1269. (13) Xu, M.; Yan, X. W.; Xie, Q. Q.; Yang, L. M.; Wang, Q. Q. Anal. Chem. 2010, 82, 1616-1620. (14) Yan, X. W.; Yang, L. M.; Wang, Q. Q. Angew. Chem. Int. Ed. 2011, 50, 5130-5133. (15) Yan, X. W.; Luo, Y. C.; Zhang, Z. B.; Li, Z. X.; Luo, Q.; Yang, L. M.; Zhang, B.; Chen, H. F.; Bai, P. M.; Wang, Q. Q. Angew. Chem. Int. Ed. 2012, 51, 3358-3363. (16) Han, G. J.; Zhang, S. C.; Xing, Z.; Zhang, X. R. Angew. Chem. Int. Ed. 2013, 52, 1466-1471.

(17) Bruckner, K.; Schwarz, K.; Beck S.; Linscheid, M. W. Anal. Chem. 2014, 86, 585-591. (18) Zhang, Z. B.; Luo, Q.; Yan, X. W.; Li, Z. X.; Luo, Y. C.; Yang, L. M.; Zhang, B.; Chen, H. F.; Wang, Q. Q. Anal. Chem. 2012, 84, 8946-8951. (19) Luo, Y. C.; Yang, X. W.; Huang, Y. S.; Wen, R. B.; Li, Z. X.; Yang, L. M.; James Yang, C. Y.; Wang, Q. Q. Anal. Chem. 2013, 85, 9428-9432. (20) Liang, Y.; Jiang, X.; Yuan, R.; Zhou, Y.; Ji, C. X.; Yang, L. M.; Chen, H. F.; Wang Q. Q. Anal. Chem. 2017, 89, 538-543. (21) Liu, C. L.; Lu, S.; Yang, L. M.; Chen, P. J.; Bai, P. M.; Wang, Q. Q. Anal. Chem. 2017, 89, 9239-9246. (22) Ornatsky, O. I.; Kinach, R.; Bandura, D. R.; Lou, X.; Tanner, S. D.; Baranov, V. I.; Nitz, M.; Winnik, M. A. J. Anal. At. Spectrom. 2008, 23, 463-469. (23) Zhao, Q., Lu, X., Yuan, C. G., Li, X. F., and Le, X. C. Anal. Chem. 2009, 81, 7484-7489. (24) Abdelrahman, A. I.; Dai, S.; Thickett, S. C.; Ornatsky, O.; Bandura, D.; Baranov, V; Winnik, M. A. J. Am. Chem. Soc. 2009, 131, 15276-15283. (25) Li, F.; Zhao, Q.; Wang, C. A.; Lu, X. F.; Li X. F.; Le, X. C. Anal. Chem. 2010, 82, 3399-3403. (26) Han, G. J.; Xing, Z.; Dong, Y. H.; Zhang, S. C.; Zhang, X. R., Angew. Chem. Int. Ed. 2011, 50 (15), 3462-3465. (27) Liu, J. M.; Yan, X. P. J. Anal. At. Spectrom. 2011, 26, 11911197. (28) Zhang, Y.; Chen, B.B.; He, M.; Yang, B.; Zhang, J.; Hu, B. Anal. Chem. 2014, 86, 8082-8089. (29) Zhang, S. X.; Han, G. J.; Xing, Z.; Zhang, S. C.; Zhang, X. R. Anal. Chem. 2014, 86, 3541-3547. (30) Yang, B.; Chen, B. B.; He, M.; Hu, B. Anal. Chem. 2017, 89, 1879-1886. (31) Liu, R.; Wang, C. Q.; Xu, Y. M.; Hu, J. Y.; Deng, D. Y.; Lv, Y. Anal. Chem. 2017, 89, 13269-13274. (32) Daniel, M. C.; Astruc, D. Chem. Rev. 2004, 104, 293-346. (33) Vancaeyzeele, C.; Ornatsky, O.; Baranov, V.; Shen, L.; Abdelrahman, A.; Winnik, M. A. J. Am. Chem. Soc. 2007, 129, 1365313660. (34) Zhai, J.; Wang, Y. L.; Xu, C.; Zheng, L. N.; Wang, M.; Feng, W. Y.; Gao, L.; Zhao, L. N.; Liu, R.; Gao, F. P.; Zhao, Y. L.; Chai, Z. F.; Gao, X. Y. Anal. Chem. 2015, 87, 2546-2549. (35) Sanchez-Rodriguez, S. P.; Munch-Anguiano, L.; BustosJaimes, I. Curr. Chem. Biol. 2010, 4, 231-243. (36) Schwarz, B.; Douglas, T. WIREs Nanomed. Nanobiotechnol. 2015, 7, 722-735. (37) Golmohammadi, R.; Valegard, K.; Fridborg, K.; Liljas, L. J. Mol. Biol. 1993, 234, 620-639. (38) Agard, N. J.; Prescher, J. A.; Bertozzi, C. R. J. Am. Chem. Soc. 2004, 126, 15046-5047. (39) Chen, C.; Ke, J.; Zhou, X. E.; Yi, W.; Brunzelle, J. S.; Li, J.; Yong, E. L.; Xu, H. E.; Melcher, K. Nature 2013, 500, 486-489. (40) Destito, G.; Yeh, R.; Rae, C. S.; Finn, M. G.; Manchester, M. Chem. Biol. 2007, 14, 1152-1162. (41) Bandura, D. R.; Baranov, V. I.; Ornatsky, O. I.; Antonov, A.; Kinach, R.; Lou, X. D.; Pavlov, S.; Vorobiev, S.; Dick, J. E.; Tanner. S. D. Anal. Chem., 2009, 81, 6813-6822. (42) Bendall1, S. C.; Simonds1, E. F.; Qiu, P.; Amir, E. D.; Krutzik, P. O.; Finck, Rachel R.; Bruggner, V.; Melamed, R.; Trejo, A.; Ornatsky, O. I.; Balderas, R. S.; Plevritis, S. K.; Sachs, K.; Pe’er, D.; Tanner, S. D. Nolan, G. P. Science 2011, 332, 687-696. (43) Han, G. J.; Spitzer, M.; Bendall, S.; Fantl, W.; Nolan, G. P. Nat. Protoc. 2018, 13, 2121-2148. (44) Strable, E.; Prasuhn, D. E.; Udit, A. K.; Brown, S.; Link, A. J.; Ngo, J. T.; Lander, G.; Quispe, J.; Potter, C. S.; Carragher, B.; Tirrell, D. A.; Finn, M. G. Bioconjugate Chem. 2008, 19, 866-875. (45) Andris Z. Mol Biotechnol 2013, 53, 92-107.


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Figure 1. Construction of Eu-tagged MS2 with folic acid targeting group. The amino group (-NH2) on MS2 was first reacted with NHS-PEGn-N3 to prepare MS2-N3 (n = 1, 4, 8 and 12, respective-ly). And then DBCOPEG-FA and DBCO-DOTA-Eu were simul-taneously modified to prepare FA-PEG-MS2-DOTA-Eu. 73x24mm (300 x 300 DPI)


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Figure 2. 153Eu-SUID-SEC/ICPMS quantification and H3[P(W3O10)4] negatively stained TEM imaging of MS2(DOTA-Eu)1034. Eu signal intensities of MS2-(DOTA-Eu)1034 and DBCO-DOTA-Eu complex (10 nM each) (a); signal intensities of 151Eu (black) and 153Eu (red) (b) and their mass flow chromatogram (c). TEM imaging (scale bar 50 nm) of MS2-N3 (d), MS2-(DOTA-Eu)1034 (e), and (FA-PEG)69-MS2-(DOTA-Eu)965 (f). 273x147mm (300 x 300 DPI)



Figure 3. Relationship of 153Eu intensity (153Euspike × 99.9% + 153EuSample × 52.19%) determined using

153Eu-SUID-ICPMS

against the number of KB cells labeled with (FA-PEG)69-MS2-(DOTA-Eu)965 (blue) and FA-PEG-DOTA-Eu (red) (n = 5). 167x64mm (300 x 300 DPI)


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TOC 176x60mm (300 x 300 DPI)


A Virus-like Element-tagged Nanoparticle ICPMS Signal Multiplier

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