On-chip sample preparation by controlled release of antibodies for simple CD4 counting

Beck, Markus; Brockhuis, Silvia; Velde, Niels van der; Breukers, Christian; Greve, Jan; Terstappen, Leon W.M.M.

doi:10.1039/c1lc20565j

Cited by 29 publications

(43 citation statements)

References 37 publications

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“…1c and 1d) is described in detail elsewhere. 17 Briefly, the gelatin/antibody-conjugate layer was cast in a chamber (15 mm × 9 mm) created by cut-outs in laminating adhesive on a substrate slide. The chamber was covered by a capping slide with openings on both ends.…”

Section: Experiments Flow Chamber and Cell Counting Chamber Fabricationmentioning

confidence: 99%

“…17 In short, our cell counting chambers contain a dry gelatin layer with embedded fluorescently labeled antibody for the immunostaining of Tlymphocytes. Inflowing blood initiates the rehydration (i.e.…”

Section: Introductionmentioning

confidence: 99%

“…16 To realize controlled reagent release, hydrogels with embedded reagents in microfluidics chambers have been employed in the past. It has been shown that sample inflow can induce swelling due to rehydration 17 or a change in pH 18 and lead to subsequent diffusion of reagents out of the hydrogel matrix. We previously demonstrated this principle for on-chip sample preparation in cell counting chambers for the enumeration of CD4-positive (CD4 + ) T-lymphocytes in a stopped-flow configuration.…”

Section: Introductionmentioning

confidence: 99%

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Controlled antibody release from gelatin for on-chip sample preparation

Zhang¹,

Wasserberg²,

Breukers³

et al. 2016

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A practical way to realize on-chip sample preparation for point-of-care diagnostics is to store the required reagents on a microfluidic device and release them in a controlled manner upon contact with the sample. For the development of such diagnostic devices, a fundamental understanding of the release kinetics of reagents from suitable materials in microfluidic chips is therefore essential. Here, we study the release kinetics of fluorophore-conjugated antibodies from (sub-) µm thick gelatin layers and several ways to control the release time. The observed antibody release is well-described by a diffusion model. Release times ranging from ~20 s to ~650 s were determined for layers with thicknesses (in the dry state) between 0.25 µm and 1.5 µm, corresponding to a diffusivity of 0.65 µm 2 /s (in the swollen state) for our standard layer preparation conditions. By modifying the preparation conditions, we can influence the properties of gelatin to realize faster or slower release. Faster drying at increased temperatures leads to shorter release times, whereas slower drying at increased humidity yields slower release. As expected in a diffusive process, the release time increases with the size of the antibody. Moreover, the ionic strength of the release medium has a significant impact on the release kinetics. Applying these findings to cell counting chambers with on-chip sample preparation, we can tune the release to control the antibody distribution after inflow of blood in order to achieve homogeneous cell staining. IntroductionOne of the major fields of applications for microfluidics is in vitro diagnostics, with great potential particularly in point-of-care diagnostics. 1-3 Many process steps and sensing principles have been realized on microfluidic devices. 4 Recently, on-chip sample preparation using reagents stored in microfluidic devices has received more and more attention. 5 On-chip sample preparation can eliminate the dependence on external instrumentation for reagent delivery as well as benchtop sample treatment, thus integrating the complete test into one disposable. 6 To realize on-chip sample preparation, reagents are integrated in microfluidic devices and released upon contact with the inflowing sample. Compared with liquid reagents 7-10 , integrating dry reagents in microfluidic devices is beneficial for long-term storage and convenient for transportation, due to the better stability of reagents in the dry state. 11 However, the controlled dissolution of the reagent and on-chip mixing with the added sample is challenging. Especially in applications where the inflowing sample passes a reservoir of the reagent and slowly dissolves or washes out the stored reagent 12-15 , very precise control of the sample flow and the reagent release process is required. In contrast, mixing between sample and reagents in stopped flow applications can be realized simply by releasing the reagent with sufficient delay after the sample inflow has stopped at the required position on the chip. 16 To realize controlled reagen...

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Section: Experiments Flow Chamber and Cell Counting Chamber Fabricationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Controlled antibody release from gelatin for on-chip sample preparation

Zhang¹,

Wasserberg²,

Breukers³

et al. 2016

Analyst

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“…67 Several of these devices use counting chambers with integrated reagents for on-chip immunostaining and a portable fluorescence imager for the detection of stained cells. 68,69 In our approach (described in details in the following chapters of this thesis) 70 , unlysed whole blood flows into the chambers driven by capillary forces, and antibodies are released into the sample after the inflow has stopped. Fluorescence imaging after 30 min incubation yields sufficient contrast between cells and background without the need for a washing step.…”

Section: On-chip Immunostaining and Cell Identificationmentioning

confidence: 99%

“…Based on this principle, we have developed a self-contained chip with on-chip sample preparation for CD4 counting. 70 One side of the interior surface in the chamber is covered with a manually deposited dry gelatin layer, which contains fluorophore-labeled antiCD3 antibody (αCD3) and antiCD4 antibody (αCD4). The chamber is filled with whole blood through capillary action.…”

Section: On-chip Immunostaining and Cell Identificationmentioning

confidence: 99%

Hands-free cytometry of whole blood : controlled antibody release from hydrogels for on-chip cell staining

Zhang¹

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Advances in Reagents Storage and Release in Self‐Contained Point‐of‐Care Devices

Deng

Jiang

2019

Adv Materials Technologies

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Microfluidics has emerged as a useful material for point‐of‐care testing. However, conventional microfluidic devices rely on multiple steps to complete the detection process, which may not be acceptable for untrained users. The emerging “self‐contained” point‐of‐care devices, with all necessary reagents prestored in, address this limitation. This paper summarizes the recent advances of self‐contained point‐of‐care devices, with emphasis on the approaches for reagents integration and manipulation. Due to the intrinsic similarity and relevance of these approaches in pharmaceutical applications, several examples about drug delivery in vivo are presented. Different approaches are compared and future trends for the development of self‐contained point‐of‐care devices are outlined.

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On-chip sample preparation by controlled release of antibodies for simple CD4 counting

Cited by 29 publications

References 37 publications

Controlled antibody release from gelatin for on-chip sample preparation

Controlled antibody release from gelatin for on-chip sample preparation

Hands-free cytometry of whole blood : controlled antibody release from hydrogels for on-chip cell staining

Advances in Reagents Storage and Release in Self‐Contained Point‐of‐Care Devices

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