Evaluation of Surface−Protein Binding Constants by Matrix-Assisted Laser Desorption Ionization Mass Spectrometry

Zhang, Ji; Kinsel, Gary R.

doi:10.1021/la026705r

Cited by 7 publications

(7 citation statements)

References 27 publications

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“…K is then calculated to be ∼7 × 10 6 M −1 . This binding constant between electrospun PCL membrane and HSA-FITC is consistent with other binding constants of proteins on hydrophobic surfaces . This confirms that electrospun PCL membranes surface are suitable for antibody immobilization via direct hydrophobic adsorption.…”

Section: Resultssupporting

confidence: 87%

Immunoassay on Free-Standing Electrospun Membranes

Han

Steckl

2009

ACS Appl. Mater. Interfaces

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For the purpose of immunoassay, electrospun membranes can be thought as the threadlike self-assembling of nano/microbeads. Nonwoven membranes of electrospun poly(epsilon-caprolactone) (PCL) fibers display excellent tenacity, flexibility and suitable surface energy. These PCL membranes exhibit easy handling in air, fast spreading, and wetting in aqueous solution, and rapid adsorption of protein molecules by hydrophobic interaction. After a fold-and-press process, the membrane porosity was reduced from approximately 75% to less than 10%, whereas the thickness increased from 5.3 to 280 microm. The resulting fluorescence signal from adsorbed protein increased>120x. With anti-HSA and HSA-FITC as an immunoassay model, a linear detection range from 500 ng/mL down to 1 ng/mL is obtained, with a detection of limit (LOD) of approximately 0.08 ng/mL. By comparison, conventional nitrocellulose and a 24.3 microm PCL fiber electrospun membrane displayed a much higher LOD of approximately 100 ng/mL. Immunoassay on free-standing electrospun membrane successfully combines the low-cost and simplicity of conventional membrane immunoassay, with the fast reaction speed and high sensitivity characteristic of magnetic nano/microbeads bioassays.

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Section: Resultssupporting

confidence: 87%

Immunoassay on Free-Standing Electrospun Membranes

Han

Steckl

2009

ACS Appl. Mater. Interfaces

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“…As mentioned above, surface MALDI-TOF measurements are difficult to quantify, and the method itself is semiquantitative. In a series of publications by the group of Kinsel, the problems to quantify this method have been discussed. − This is very difficult especially for a very little amount of adsorbed protein. However, the detection limits of the method are competitive, and surface MALDI-TOF methods are recently becoming more and more important …”

Section: Resultsmentioning

confidence: 99%

Comparison of Coatings from Reactive Star Shaped PEG-stat-PPG Prepolymers and Grafted Linear PEG for Biological and Medical Applications

et al. 2005

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Grafting of poly(ethylene glycol) (PEG) is a common strategy for reducing nonspecific interactions of surfaces with proteins. We have used grafting at "cloud point" solution conditions that ensures maximum grafting density of linear methoxy terminated PEG-aldehyde (mPEG-ald, M(w) = 5000 and 30000). In an alternative approach, surfaces were modified with layers prepared from isocyanate terminated, star shaped poly(ethylene glycol-stat-propylene glycol) prepolymers (80% ethylene glycol, six arms, M(w) = 3000, 12,000, and 18,000; this compound will be referred to as "Star PEG" in the text). Due to the highly reactive endgroups, these molecules form a dense network on the substrate with a high polymer surface coverage. The two systems were compared regarding their ability to prevent unspecific adsorption of insulin and lysozyme. The layers were analyzed by ellipsometry, contact angle measurements, and XPS. Protein adsorption was monitored by surface MALDI-TOF MS and fluorescence microscopy. No protein adsorption could be detected on Star PEG coatings and on mPEG-ald 5000, whereas mPEG-ald 30,000 could only prevent adsorption of lysozyme but not of the smaller insulin.

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“…Furthermore, the number of desorbed ions that do form by energetic desorption methods often fluctuates dramatically due to the competition of different ionization mechanisms and their dependencies upon diverse surface properties. For example, the MALDI signal for a given species is generally found to be inversely proportional to its surface binding strength, inhibiting detection of covalently and other strongly bound species. , Such fluctuations in ion yields complicate identification and quantification of organic surface species, limiting the applicability of these techniques to surface analysis.…”

mentioning

confidence: 99%

Derivatization of Surface-Bound Peptides for Mass Spectrometric Detection via Threshold Single Photon Ionization

et al. 2004

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Chemical derivatization of peptides allows efficient F2 laser single photon ionization (SPI) of Fmoc-derivatized peptides covalently bound to surfaces. Laser desorption photoionization mass spectrometry using 337-nm pulses for desorption and 157.6-nm pulses for threshold SPI forms large ions identified as common peptide fragments bound to either Fmoc or the surface linker. Electronic structure calculations indicate the Fmoc label is behaving as an ionization tag for the entire peptide, lowering the ionization potential of the complex below the 7.87-eV photon energy. This method should allow detection of many molecular species covalently or electrostatically bound to surfaces.

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Evaluation of Surface−Protein Binding Constants by Matrix-Assisted Laser Desorption Ionization Mass Spectrometry

Cited by 7 publications

References 27 publications

Immunoassay on Free-Standing Electrospun Membranes

Immunoassay on Free-Standing Electrospun Membranes

Comparison of Coatings from Reactive Star Shaped PEG-stat-PPG Prepolymers and Grafted Linear PEG for Biological and Medical Applications

Derivatization of Surface-Bound Peptides for Mass Spectrometric Detection via Threshold Single Photon Ionization

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