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A disposable, self-contained PCR chip.

ABSTRACT: A disposable, self-contained polymerase chain reaction (PCR) chip with on-board stored, just-on-time releasable, paraffin-passivated, dry reagents is described. During both storage and sample preparation, the paraffin immobilizes and protects the stored reagents. Fluid flow through the reactor leaves the reagents undisturbed. Prior to the amplification step, the chamber is filled with target analyte suspended in water. Upon heating the PCR chamber to the DNA's denaturation temperature, the paraffin melts and moves out of the way, and the reagents are released and hydrated. To better understand the reagent release process, a scaled up model of the reactor was constructed and the paraffin migration was visualized. Experiments were carried out with a 30 microl reactor demonstrating detectable amplification (with agarose gel electrophoresis) of 10 fg ( approximately 200 copies) of lambda DNA template. The in-reactor storage and on-time release of the PCR reagents reduce the number of needed operations and significantly simplifies the flow control that would, otherwise, be needed in lab-on-chip devices.

SUBMITTER: Kim J

PROVIDER: S-EPMC2676225 | biostudies-literature | 2009 Feb

REPOSITORIES: biostudies-literature

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A disposable, self-contained PCR chip.

Kim Jitae J Byun Doyoung D Mauk Michael G MG Bau Haim H HH

Lab on a chip 20081118 4

A disposable, self-contained polymerase chain reaction (PCR) chip with on-board stored, just-on-time releasable, paraffin-passivated, dry reagents is described. During both storage and sample preparation, the paraffin immobilizes and protects the stored reagents. Fluid flow through the reactor leaves the reagents undisturbed. Prior to the amplification step, the chamber is filled with target analyte suspended in water. Upon heating the PCR chamber to the DNA's denaturation temperature, the paraf ...[more]

PMID: 19190797

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Project description:Biophysical properties are widely used to detect pathophysiological processes of vascular diseases or clinical states. For early detection of cardiovascular diseases, it is necessary to simultaneously measure multiple biophysical properties in a microfluidic environment. However, a microfluidic-based technique for measuring multiple biophysical properties has not been demonstrated. In this study, a simple measurement method was suggested to quantify three biophysical properties of blood, including red blood cell (RBC) deformability, RBC aggregation, and hematocrit. To demonstrate the suggested method, a microfluidic device was constructed, being composed of a big-sized channel (BC), a parallel micropillar (MP), a main channel, a branch channel, inlet, and outlets. By operating a single syringe pump, blood was supplied into the inlet of the microfluidic device, at a periodic on-off profile (i.e., period = 240 s). The RBC deformability index (DI) was obtained by analyzing the averaged blood velocity in the branch channel. Additionally, the RBC aggregation index (AIN) and the hematocrit index (HiBC) were measured by analyzing the image intensity of blood flows in the MP and the BC, respectively. The corresponding contributions of three influencing factors, including the turn-on time (Ton), the amplitude of blood flow rate (Q₀), and the hematocrit (Hct) on the biophysical indices (DI, AIN, and HiBC) were evaluated quantitatively. As the three biophysical indices varied significantly with respect to the three factors, the following conditions (i.e., Ton = 210 s, Q₀ = 1 mL/h, and Hct = 50%) were maintained for consistent measurement of biophysical properties. The proposed method was employed to detect variations of biophysical properties depending on the concentrations of autologous plasma, homogeneous hardened RBCs, and heterogeneous hardened RBCs. Based on the observations, the proposed method exhibited significant differences in biophysical properties depending on base solutions, homogeneous hardened RBCs (i.e., all RBCs fixed with the same concentration of glutaraldehyde solution), and heterogeneous hardened RBCs (i.e., partially mixed with normal RBCs and homogeneous hardened RBCs). Additionally, the suggested indices (i.e., DI, AIN, and HiBC) were effectively employed to quantify three biophysical properties, including RBC deformability, RBC aggregation, and hematocrit.

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A disposable, self-contained PCR chip.

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A disposable, self-contained PCR chip.

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