Optimization of an Anti-poly(ethylene glycol) (anti-PEG) Cell-Based Capture System To Quantify PEG and PEGylated Molecules

Wen Wei Lin, Yuan Chin Hsieh, Yi-An Cheng, Kuo-Hsiang Chuang, Chien Chiao Huang, Chih Hung Chuang, I. Ju Chen, Kai Wen Cheng, Yun-Chi Lu, Ta Chun Cheng, Yeng Tseng Wang, Steve R. Roffler, Tian Lu Cheng

Research output: Contribution to journalArticle

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Abstract

Sensitive determination of the pharmacokinetics of PEGylated molecules can accelerate the process of drug development. Here, we combined different anti-PEG Fab expressing 293T cells as capture cells (293T/3.3, 293T/6.3, and 293T/15-2b cells) with four detective anti-PEG antibodies (3.3, 6.3, 7A4, or 15-2b) to optimize an anti-PEG cell-based sandwich ELISA. Then, we quantified free PEG (mPEG2K-NH2 and mPEG5K-NH2) or PEG-conjugated small molecules (mPEG5K-biotin and mPEG5K-NIR797), proteins (PegIntron and Pegasys), and nanoparticles (Liposomal-Doxorubicin and quantum-dots). The combination of 293T/15-2b cells and the 7A4 detection antibody was best sensitivity for free PEG, PEG-like molecules, and PEGylated proteins with detection at ng mL(-1) levels. On the other hand, 293T/3.3 cells combined with the 15-2b antibody had the highest sensitivity for quantifying Lipo-Dox at 2 ng mL(-1). All three types of anti-PEG cells combined with the 15-2b antibody had high sensitivity for quantum dot quantification down to 7 pM. These results suggest that the combination of 293T/15-2b cells and 7A4 detection antibody is the optimal pair for sensitive quantification of free PEG, PEG-like molecules, and PEGylated proteins, whereas the 293T/3.3 cells combined with 15-2b are more suitable for quantifying PEGylated nanoparticles. The optimized anti-PEG cell-based sandwich ELISA can provide a sensitive, precise, and convenient tool for the quantification of a range of PEGylated molecules.

Original languageEnglish
Pages (from-to)12371-12379
Number of pages9
JournalAnalytical Chemistry
Volume88
Issue number24
DOIs
Publication statusPublished - Dec 20 2016

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Polyethylene glycols
Molecules
Antibodies
Semiconductor quantum dots
Nanoparticles
Proteins
Pharmacokinetics
Biotin

Keywords

  • Journal Article

Cite this

Optimization of an Anti-poly(ethylene glycol) (anti-PEG) Cell-Based Capture System To Quantify PEG and PEGylated Molecules. / Lin, Wen Wei; Hsieh, Yuan Chin; Cheng, Yi-An; Chuang, Kuo-Hsiang; Huang, Chien Chiao; Chuang, Chih Hung; Chen, I. Ju; Cheng, Kai Wen; Lu, Yun-Chi; Cheng, Ta Chun; Wang, Yeng Tseng; Roffler, Steve R.; Cheng, Tian Lu.

In: Analytical Chemistry, Vol. 88, No. 24, 20.12.2016, p. 12371-12379.

Research output: Contribution to journalArticle

Lin, WW, Hsieh, YC, Cheng, Y-A, Chuang, K-H, Huang, CC, Chuang, CH, Chen, IJ, Cheng, KW, Lu, Y-C, Cheng, TC, Wang, YT, Roffler, SR & Cheng, TL 2016, 'Optimization of an Anti-poly(ethylene glycol) (anti-PEG) Cell-Based Capture System To Quantify PEG and PEGylated Molecules', Analytical Chemistry, vol. 88, no. 24, pp. 12371-12379. https://doi.org/10.1021/acs.analchem.6b03614
Lin, Wen Wei ; Hsieh, Yuan Chin ; Cheng, Yi-An ; Chuang, Kuo-Hsiang ; Huang, Chien Chiao ; Chuang, Chih Hung ; Chen, I. Ju ; Cheng, Kai Wen ; Lu, Yun-Chi ; Cheng, Ta Chun ; Wang, Yeng Tseng ; Roffler, Steve R. ; Cheng, Tian Lu. / Optimization of an Anti-poly(ethylene glycol) (anti-PEG) Cell-Based Capture System To Quantify PEG and PEGylated Molecules. In: Analytical Chemistry. 2016 ; Vol. 88, No. 24. pp. 12371-12379.
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T1 - Optimization of an Anti-poly(ethylene glycol) (anti-PEG) Cell-Based Capture System To Quantify PEG and PEGylated Molecules

AU - Lin, Wen Wei

AU - Hsieh, Yuan Chin

AU - Cheng, Yi-An

AU - Chuang, Kuo-Hsiang

AU - Huang, Chien Chiao

AU - Chuang, Chih Hung

AU - Chen, I. Ju

AU - Cheng, Kai Wen

AU - Lu, Yun-Chi

AU - Cheng, Ta Chun

AU - Wang, Yeng Tseng

AU - Roffler, Steve R.

AU - Cheng, Tian Lu

PY - 2016/12/20

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N2 - Sensitive determination of the pharmacokinetics of PEGylated molecules can accelerate the process of drug development. Here, we combined different anti-PEG Fab expressing 293T cells as capture cells (293T/3.3, 293T/6.3, and 293T/15-2b cells) with four detective anti-PEG antibodies (3.3, 6.3, 7A4, or 15-2b) to optimize an anti-PEG cell-based sandwich ELISA. Then, we quantified free PEG (mPEG2K-NH2 and mPEG5K-NH2) or PEG-conjugated small molecules (mPEG5K-biotin and mPEG5K-NIR797), proteins (PegIntron and Pegasys), and nanoparticles (Liposomal-Doxorubicin and quantum-dots). The combination of 293T/15-2b cells and the 7A4 detection antibody was best sensitivity for free PEG, PEG-like molecules, and PEGylated proteins with detection at ng mL(-1) levels. On the other hand, 293T/3.3 cells combined with the 15-2b antibody had the highest sensitivity for quantifying Lipo-Dox at 2 ng mL(-1). All three types of anti-PEG cells combined with the 15-2b antibody had high sensitivity for quantum dot quantification down to 7 pM. These results suggest that the combination of 293T/15-2b cells and 7A4 detection antibody is the optimal pair for sensitive quantification of free PEG, PEG-like molecules, and PEGylated proteins, whereas the 293T/3.3 cells combined with 15-2b are more suitable for quantifying PEGylated nanoparticles. The optimized anti-PEG cell-based sandwich ELISA can provide a sensitive, precise, and convenient tool for the quantification of a range of PEGylated molecules.

AB - Sensitive determination of the pharmacokinetics of PEGylated molecules can accelerate the process of drug development. Here, we combined different anti-PEG Fab expressing 293T cells as capture cells (293T/3.3, 293T/6.3, and 293T/15-2b cells) with four detective anti-PEG antibodies (3.3, 6.3, 7A4, or 15-2b) to optimize an anti-PEG cell-based sandwich ELISA. Then, we quantified free PEG (mPEG2K-NH2 and mPEG5K-NH2) or PEG-conjugated small molecules (mPEG5K-biotin and mPEG5K-NIR797), proteins (PegIntron and Pegasys), and nanoparticles (Liposomal-Doxorubicin and quantum-dots). The combination of 293T/15-2b cells and the 7A4 detection antibody was best sensitivity for free PEG, PEG-like molecules, and PEGylated proteins with detection at ng mL(-1) levels. On the other hand, 293T/3.3 cells combined with the 15-2b antibody had the highest sensitivity for quantifying Lipo-Dox at 2 ng mL(-1). All three types of anti-PEG cells combined with the 15-2b antibody had high sensitivity for quantum dot quantification down to 7 pM. These results suggest that the combination of 293T/15-2b cells and 7A4 detection antibody is the optimal pair for sensitive quantification of free PEG, PEG-like molecules, and PEGylated proteins, whereas the 293T/3.3 cells combined with 15-2b are more suitable for quantifying PEGylated nanoparticles. The optimized anti-PEG cell-based sandwich ELISA can provide a sensitive, precise, and convenient tool for the quantification of a range of PEGylated molecules.

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DO - 10.1021/acs.analchem.6b03614

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VL - 88

SP - 12371

EP - 12379

JO - Analytical Chemistry

JF - Analytical Chemistry

SN - 0003-2700

IS - 24

ER -