研究目的
To identify a novel non-antibody-binding peptide (nABP296) with high affinity for CD105 for use in tumour imaging and diagnosis, specifically for osteosarcoma.
研究成果
nABP296 is a novel peptide that specifically binds to CD105, enabling effective imaging of osteosarcoma in vitro and in vivo. It shows potential for diagnosis, MSC isolation, and targeted therapy, with no cytotoxicity observed.
研究不足
The mechanism underlying the penetrating effect of nABP296 is unclear and needs further investigation. The study focused on specific cell lines and models, and generalizability to other cancers or in vivo human applications may be limited.
1:Experimental Design and Method Selection:
Used M13 phage biopanning to identify peptides binding to CD105, followed by ELISA, flow cytometry, immunofluorescence, and in vivo imaging to confirm binding affinity and specificity.
2:Sample Selection and Data Sources:
Used MNNG/HOS (CD105-positive) and Cal27 (CD105-negative) cell lines, recombinant human CD105 protein, xenograft tumour models in mice, and human osteosarcoma and tongue carcinoma tissue sections.
3:List of Experimental Equipment and Materials:
Included M13 phage display library, MaxiSorp plates, flow cytometer, confocal laser scanning microscope, IVIS Spectrum In Vivo Imaging system, and various antibodies and reagents.
4:Experimental Procedures and Operational Workflow:
Steps involved biopanning, peptide synthesis, cell culture, RT-PCR, immunofluorescence, flow cytometry, ELISA, cytotoxicity assays, in vivo imaging, and histological staining.
5:Data Analysis Methods:
Statistical analysis using GraphPad Prism software, with one-way ANOVA for cytotoxicity data.
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Confocal laser scanning microscopy
CLSM
Leica Microsystems GmbH
Used for immunofluorescence imaging to visualize peptide and antibody binding.
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IVIS Spectrum In Vivo Imaging system
IVIS Spectrum
PerkinElmer, Inc.
Used for in vivo imaging to detect fluorescence from labelled peptides in tumour-bearing mice.
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VICTOR X5 Multilabel plate reader
VICTOR X5
PerkinElmer, Inc.
Used for measuring absorbance in ELISA and cytotoxicity assays.
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Ph.D.-12 Phage Display Peptide Library
Ph.D.-12
New England Biolabs, Inc.
Used for biopanning to identify peptides binding to CD105.
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MaxiSorp plates
MaxiSorp
Thermo Fisher Scientific, Inc.
Used for coating proteins in ELISA and biopanning experiments.
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FACSCalibur flow cytometer
FACSCalibur
BD Biosciences
Used for flow cytometry analysis to measure CD105 expression and peptide binding.
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NanoDrop 1000
NanoDrop 1000
Thermo Fisher Scientific, Inc.
Used for spectrophotometry to measure RNA concentration.
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Cell Counting Kit-8
CCK-8
Dojindo Molecular Technologies, Inc.
Used for cytotoxicity assays to measure cell viability.
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Microm HM560 cryostat microtome
Microm HM560
Thermo Fisher Scientific, Inc.
Used for sectioning tumour tissues for histological analysis.
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