Paper-based immunocapture for targeted protein analysis

Skjærvø, Øystein; Solbakk, Eirik J.; Halvorsen, Trine Grønhaug; Reubsaet, Léon

doi:10.1016/j.talanta.2018.12.013

Cited by 15 publications

(21 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Functionalization using HEMA-VDM: For the HEMA-VDM functionalization, the procedure published by Skjaervø et al [29,[36][37][38] was slightly modified and used. In short, the fibers on the paper discs were first silanized by applying 10 µL of freshly prepared silanizing solution (this is a mixture of 2.5 mg DPPH, 330 mg DMF and 172 mg γ-MAPS prepared in a glass vial).…”

Section: Functionalization Of Paper Discs and Protein Immobilizationmentioning

confidence: 99%

“…An easy way to determine if there is protein bound to the paper is by CBB staining. To do this for our purpose, a previously described staining procedure for paper was used [37,39]. It was expected that the protein immobilized on untreated paper followed by a wash would yield minimally stained discs, while protein immobilization on functionalized paper followed by a wash would yield strongly stained discs.…”

Section: Staining Of Proteins On Paper To Assess Protein Bindingmentioning

confidence: 99%

“…The long-term goal of our research is to simplify MS-based protein analysis from dried matrix samples (such as DBS), and we aim to integrate time-consuming procedures within the sampling device for a prompt start to the sample preparation upon sample collection [29,[35][36][37][38].…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper

et al. 2021

Self Cite

View full text Add to dashboard Cite

This paper describes smart sampling paper to be used for bottom-up protein analysis. Four different manners to immobilize trypsin on cellulose were evaluated. Untreated paper, potassium-periodate-functionalized paper (with and without post-immobilization reduction) and 2-hydroxyethyl methacrylate (HEMA)/2-vinyl-4,4-dimethylazlactone (VDM)-functionalized paper were all used to immobilize trypsin. For the evaluation, Coomassie Brilliant Blue staining of proteins on paper and the BAEE trypsin activity assay needed to be modified. These methods allowed, together with data from mass spectrometric analysis of cytochrome C digestions, us to acquire fundamental insight into protein binding, and trypsin action and activity on paper. All functionalized discs bind more protein than the untreated discs. Protein binding to functionalized discs is based on both adsorption and covalent binding. Trypsin immobilized on potassium-periodate-functionalized discs exhibits the highest trypsin activity when using cytochrome C as substrate. It is proven that it is trypsin attached to paper (and not desorbed trypsin) which is responsible for the enzyme activity. The use of discs on complex biological samples shows that all functionalized discs are able to digest diluted serum; for the best-performing disc, HEMA-VDM functionalized, up to 200 high-confidence proteins are qualified, showing its potential.

show abstract

Section: Functionalization Of Paper Discs and Protein Immobilizationmentioning

confidence: 99%

Section: Staining Of Proteins On Paper To Assess Protein Bindingmentioning

confidence: 99%

See 1 more Smart Citation

Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper

et al. 2021

Self Cite

View full text Add to dashboard Cite

show abstract

“…Our group has been working with the concept of smart sampling in combination with MS determination since 2017 [45]. This concept is based on the covalent binding of trypsin to cellulose allowing a DBS sampler to start with the digestion step at the moment of sampling [46][47][48][49]. This smart sampler does not compromise the ease of sampling as the format remains practically unchanged.…”

Section: Introductionmentioning

confidence: 99%

Smart proteolysis samplers for pre‐lab bottom‐up protein analysis – Performance of on‐paper digestion compared to conventional digestion

Nguyen

Halvorsen

Thiede

et al. 2022

Separation Science Plus

Self Cite

View full text Add to dashboard Cite

Here the relation between digestion of proteins by trypsin covalently bound to paper and trypsin in‐solution is investigated. The trypsin acting on paper is covalently bound. A trypsin concentration of 0.5% (w/v) results in the highest digestion activity of all concentrations tested. Additionally, it can be seen that trypsin on‐paper has retained approx. 50% of its activity. Unlike trypsin in‐solution, the stability of the smart proteolysis samplers was regarded to be stable for at least four months when kept refrigerated. Autolysis was very small for covalently bound trypsin: less than 2% compared to in‐solution trypsin. Proteomic analysis of diluted human serum showed more protein identifications (214) in‐solution digestions than on‐paper digestions (76). Also, higher coverage for the in‐solution digestion was obtained. Those proteins identified after on‐paper digestion with no or few disulfide bonds seem to have more similar sequence coverages compared to those identified after in‐solution digestion. Smart samplers allow the determination of at least 70–75 proteins without performing the overnight digestion. All in all, trypsin covalently bound to paper shows to retain high proteolytic activity and is a stable alternative for conventional digestions. In this way, smart proteolytic samplers show their feasibility in pre‐lab sample preparation.

show abstract

“…These smart samplers allow the pretreatment to start already upon blood collection, thus saving valuable time and material in the preparation of the sample later on. Integration of sample pretreatment steps as, for example, tryptic digestion for bottom-up protein analysis [ 16 – 18 ] or affinity capture for sample clean-up [ 19 , 20 ] on, for instance, cellulose [ 21 ] is feasible. Technically, biomolecules such as proteins can be immobilized on paper in three different ways: (1) through adsorption due to weak, non-covalent, intermolecular forces; (2) coupling via a bioaffinity tag; or (3) linked through covalent binding.…”

Section: Introductionmentioning

confidence: 99%

On the spot immunocapture in targeted biomarker analysis using paper-bound streptavidin as anchor for biotinylated antibodies

et al. 2022

Self Cite

View full text Add to dashboard Cite

The modification of an easily available resource like paper to circumvent expensive or intensive sample pretreatment could be the answer to sample analysis in resource-poor regions. Therefore, a novel on-paper device combining sample collection with affinity sample pretreatment is introduced here. Universal smart affinity samplers are produced by a simple KIO4-mediated oxidation of cellulose, which functionalizes the paper. This is followed by immobilization of streptavidin. Streptavidin serves as a universal anchor for biotinylated antibodies, enabling simple preparation of tailor-made affinity samplers. The functionality of the device was tested using a model protein (human chorionic gonadotropin, hCG) and biotinylated anti-hCG antibodies for affinity capture. In a laboratory setting, the performance was demonstrated, and a 14-fold increase of target binding compared to binding without bmAb was achieved. The recovery of hCG captured with bmAb-treated samplers was determined to be 33% and comparable to previously described affinity capture approaches. Application of the smart affinity samplers to human serum containing hCG showed an R2 of 0.98 (200–1000 pg mL−1), precision of ≤ 9.1% RSD, and estimated limit of detection of 65 pg mL−1. Although further optimization and validation are necessary prior to application to real samples in clinical settings, the potential of the device for use in determination of low abundant biomarkers in complex samples has been demonstrated.

show abstract

Paper-based immunocapture for targeted protein analysis

Cited by 15 publications

References 28 publications

Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper

Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper

Smart proteolysis samplers for pre‐lab bottom‐up protein analysis – Performance of on‐paper digestion compared to conventional digestion

On the spot immunocapture in targeted biomarker analysis using paper-bound streptavidin as anchor for biotinylated antibodies

Contact Info

Product

Resources

About