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A new strategy to screen molecular imaging probe uptake in cell culture without radiolabeling using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
J Nucl Med. 2005 May; 46(5):878-86.JN

Abstract

Numerous new molecular targets for diseases are rapidly being identified and validated in the postgenomic era, urging scientists to explore novel techniques for accelerating molecular probe development. In this study, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was investigated as a potential tool for high-throughput screening and characterization of molecular imaging probes. Specifically, MALDI-TOF-MS was used to screen a small library of phosphonium cations for their ability to accumulate in cells.

METHODS

C6 cells incubated with phosphonium cations at room temperature were collected and lysed for experiments. Calibration curves for the internal standard, methyltriphenyl phosphonium, and for tetraphenylphosphonium bromide (TPP) and other phosphonium cations were first established. The time course of TPP uptake by C6 cells was then quantified using both MALDI-TOF-MS and liquid scintillation counting with (3)H-TPP. In addition, MALDI-TOF-MS was used to screen a library of 8 phosphonium cations and subsequently rank their ability to penetrate membranes and accumulate in cells. Finally, the accumulation of 4-fluorophenyltriphenyl phosphonium (FTPP) in the membrane potential-modulated cells was also measured by MALDI-TOF-MS.

RESULTS

MALDI-TOF-MS spectra clearly revealed that TPP was easily identified from cell lysates even as early as 10 min after incubation and that levels as low as 0.11 fmol of TPP per cell could be detected, suggesting the high sensitivity of this technique. The time course of TPP influx determined by both MALDI-TOF-MS and radioactivity counting showed no statistically significant difference (P > 0.05 for all time points). These data validated MALDI-TOF-MS as an alternative approach for accurately measuring uptake of phosphonium cations by cells. TPP and FTPP demonstrated greater accumulation in cells than did the other cations evaluated in this study. Furthermore, uptake profiles suggested that FTPP preserves the membrane potential-dependent uptake property of TPP in cell cultures. Taken together, these data justify further synthesis and evaluation of (18)F-FTPP as a molecular probe for imaging mitochondrial dysfunction.

CONCLUSION

These results demonstrate that MALDI-TOF-MS is a powerful analytic tool for rapid screening and characterization of phosphonium cations as molecular probes. This technique can potentially be applied to the evaluation of other imaging probes or drugs and thus may facilitate their rational design and development.

Authors+Show Affiliations

Molecular Imaging Program at Stanford, Department of Radiology, and Bio-X Program, Stanford University, 318 Campus Drive, Clark Center E-150, Stanford, CA 94305, USA. sgambhir@stanford.eduNo affiliation info availableNo affiliation info available

Pub Type(s)

Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

15872363

Citation

Cheng, Zhen, et al. "A New Strategy to Screen Molecular Imaging Probe Uptake in Cell Culture Without Radiolabeling Using Matrix-assisted Laser Desorption/ionization Time-of-flight Mass Spectrometry." Journal of Nuclear Medicine : Official Publication, Society of Nuclear Medicine, vol. 46, no. 5, 2005, pp. 878-86.
Cheng Z, Winant RC, Gambhir SS. A new strategy to screen molecular imaging probe uptake in cell culture without radiolabeling using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. J Nucl Med. 2005;46(5):878-86.
Cheng, Z., Winant, R. C., & Gambhir, S. S. (2005). A new strategy to screen molecular imaging probe uptake in cell culture without radiolabeling using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Journal of Nuclear Medicine : Official Publication, Society of Nuclear Medicine, 46(5), 878-86.
Cheng Z, Winant RC, Gambhir SS. A New Strategy to Screen Molecular Imaging Probe Uptake in Cell Culture Without Radiolabeling Using Matrix-assisted Laser Desorption/ionization Time-of-flight Mass Spectrometry. J Nucl Med. 2005;46(5):878-86. PubMed PMID: 15872363.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - A new strategy to screen molecular imaging probe uptake in cell culture without radiolabeling using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AU - Cheng,Zhen, AU - Winant,Richard C, AU - Gambhir,Sanjiv S, PY - 2005/5/6/pubmed PY - 2005/6/25/medline PY - 2005/5/6/entrez SP - 878 EP - 86 JF - Journal of nuclear medicine : official publication, Society of Nuclear Medicine JO - J Nucl Med VL - 46 IS - 5 N2 - UNLABELLED: Numerous new molecular targets for diseases are rapidly being identified and validated in the postgenomic era, urging scientists to explore novel techniques for accelerating molecular probe development. In this study, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was investigated as a potential tool for high-throughput screening and characterization of molecular imaging probes. Specifically, MALDI-TOF-MS was used to screen a small library of phosphonium cations for their ability to accumulate in cells. METHODS: C6 cells incubated with phosphonium cations at room temperature were collected and lysed for experiments. Calibration curves for the internal standard, methyltriphenyl phosphonium, and for tetraphenylphosphonium bromide (TPP) and other phosphonium cations were first established. The time course of TPP uptake by C6 cells was then quantified using both MALDI-TOF-MS and liquid scintillation counting with (3)H-TPP. In addition, MALDI-TOF-MS was used to screen a library of 8 phosphonium cations and subsequently rank their ability to penetrate membranes and accumulate in cells. Finally, the accumulation of 4-fluorophenyltriphenyl phosphonium (FTPP) in the membrane potential-modulated cells was also measured by MALDI-TOF-MS. RESULTS: MALDI-TOF-MS spectra clearly revealed that TPP was easily identified from cell lysates even as early as 10 min after incubation and that levels as low as 0.11 fmol of TPP per cell could be detected, suggesting the high sensitivity of this technique. The time course of TPP influx determined by both MALDI-TOF-MS and radioactivity counting showed no statistically significant difference (P > 0.05 for all time points). These data validated MALDI-TOF-MS as an alternative approach for accurately measuring uptake of phosphonium cations by cells. TPP and FTPP demonstrated greater accumulation in cells than did the other cations evaluated in this study. Furthermore, uptake profiles suggested that FTPP preserves the membrane potential-dependent uptake property of TPP in cell cultures. Taken together, these data justify further synthesis and evaluation of (18)F-FTPP as a molecular probe for imaging mitochondrial dysfunction. CONCLUSION: These results demonstrate that MALDI-TOF-MS is a powerful analytic tool for rapid screening and characterization of phosphonium cations as molecular probes. This technique can potentially be applied to the evaluation of other imaging probes or drugs and thus may facilitate their rational design and development. SN - 0161-5505 UR - https://www.unboundmedicine.com/medline/citation/15872363/A_new_strategy_to_screen_molecular_imaging_probe_uptake_in_cell_culture_without_radiolabeling_using_matrix_assisted_laser_desorption/ionization_time_of_flight_mass_spectrometry_ L2 - http://jnm.snmjournals.org/cgi/pmidlookup?view=long&pmid=15872363 DB - PRIME DP - Unbound Medicine ER -