Direct immobilization of DNA probes on non-modified plastics by UV irradiation and integration in microfluidic devices for rapid bioassay

DNA microarrays have become one of the most powerful tools in the field of genomics and medical diagnosis. Recently, there has been increased interest in combining microfluidics with microarrays since this approach offers advantages in terms of portability, reduced analysis time, low consumption of reagents, and increased system integration. Polymers are widely used for microfluidic systems, but fabrication of microarrays on such materials often requires complicated chemical surface modifications, which hinders the integration of microarrays into microfluidic systems. In this paper, we demonstrate that simple UV irradiation can be used to directly immobilize poly(T)poly(C)-tagged DNA oligonucleotide probes on many different types of plastics without any surface modification. On average, five- and fourfold improvement in immobilization and hybridization efficiency have been achieved compared to surface-modified slides with aminated DNA probes. Moreover, the TC tag only costs 30% of the commonly used amino group modifications. Using this microarray fabrication technique, a portable cyclic olefin copolymer biochip containing eight individually addressable microfluidic channels was developed and used for rapid and parallel identification of Avian Influenza Virus by DNA hybridization. The one-step, cost-effective DNA-linking method on non-modified polymers significantly simplifies microarray fabrication procedures and permits great flexibility to plastic material selection, thus making it convenient to integrate microarrays into plastic microfluidic systems

Synergism between particle-based multiplexing and microfluidics technologies may bring diagnostics closer to the patient

In the field of medical diagnostics there is a growing need for inexpensive, accurate, and quick high-throughput assays. On the one hand, recent progress in microfluidics technologies is expected to strongly support the development of miniaturized analytical devices, which will speed up (bio)analytical assays. On the other hand, a higher throughput can be obtained by the simultaneous screening of one sample for multiple targets (multiplexing) by means of encoded particle-based assays. Multiplexing at the macro level is now common in research labs and is expected to become part of clinical diagnostics. This review aims to debate on the “added value” we can expect from (bio)analysis with particles in microfluidic devices. Technologies to (a) decode, (b) analyze, and (c) manipulate the particles are described. Special emphasis is placed on the challenges of integrating currently existing detection platforms for encoded microparticles into microdevices and on promising microtechnologies that could be used to down-scale the detection units in order to obtain compact miniaturized particle-based multiplexing platforms

Three-dimensional biologically-active nanostructured architectures in polymer-based microfluidic devices for biomarker detection

We report a novel three-dimensional (3D) DNA analysis device consisting of ultrahigh-aspect-ratio polymer nanopillars prepared by a templating polymerization patterntransfer method followed by a direct-write photochemical patterning approach to produce functional scaffolds for the immobilization of molecular recognition probes to nanopillars. These nanopillars can be integrated into microfluidic channels for producing high-surface-area supports for these probes to increase assay sensitivity and dynamic range, as well as rapid response times due to microfluidic addressing of the array. Anodic aluminum oxide (AAO) was used as the template for the nanostructures as well as the molding tool for the microfluidic network. The AAO template consisted of nanopores with a close-packed hexagonal geometry. The device demonstrated highspeed hybridization kinetics with detection limits on the order of femtomoles. © 2006 Society for Chemistry and Micro-Nano Systems

Functional template-derived poly(methyl methacrylate) nanopillars for solid-phase biological reactions

A free-standing erect polymethyl methacrylate (PMMA) nanopillar that possess ultrahigh aspect ratio was fabricated with nondestructive surface functionalization technique for solid-phase biological reactions. Photopolymerization of monomer within nanometer-diameter, micrometer-tall pores of anodic aluminum oxide (AAO) templates led to the facile production of PMMA nanostructures that retain the shape and size of the template pores. The electron micrograph of PMMA pillars modified for 20 min using a direct-write UV photochemical patterning method consists of oxygen and carbon. Scanning Electron Microscopy (SEM) images for smallest nanopillar features size fabricated show no damage for exposure times of 20 min and also the production of maximum carboxylic acid surface density. The results show that the amount of substrate processes by enzymes immobilized to a surface with a fixed two-dimensional footprint can be increased by the use of PMMA nanopillar supports

Fabrication of DNA microarrays onto poly(methyl methacrylate) with ultraviolet patterning and microfluidics for the detection of low-abundant point mutations

We have developed a simple ultraviolet (UV)-photomodification protocol using poly(methyl methacrylate) and polycarbonate to produce functional scaffolds consisting of carboxylic groups that allow covalent attachment of amine-terminated oligonucleotide probes to these surface groups through carbodiimide coupling. Use of the photomodification procedure coupled to microfluidics allowed for the rapid generation of medium-density DNA microarrays. The method reported herein involves the use of poly(dimethylsiloxane) microchannels reversibly sealed to photomodified poly(methyl methacrylate) surfaces to serve as stencils for patterning the oligonucleotide probes. After array construction, the poly(dimethylsiloxane) stencil is rotated 90°to allow interrogation of the array using microfluidics. The photomodification process for array fabrication involves only three steps: (1) broadband UV exposure of the polymer surface, (2) carbodiimide coupling of amine-terminated oligonucleotide probes to the surface (via an amide bond), and (3) washing of the surface. The density of probes attached to this activated surface was found to be ∼41 pmol cm-2, near the steric-saturation limit for short oligonucleotide probes. We demonstrate the use of this procedure for screening multiple KRAS2 mutations possessing high diagnostic value for colorectal cancers. A ligase detection reaction/universal array assay was carried out using parallel detection of two different low-abundant DNA point mutations in KRAS2 oncogenes with the allelic composition evaluated at one locus. Four zip code probes immobilized onto the poly(methyl methacrylate) surface directed allele-specific ligation products containing mutations in the KRAS2 gene (12.2D, 12.2A, 12.2V, and 13.4D) to the appropriate address of a universal array with minimal amounts of cross-hybridization or misligation. © 2005 Elsevier Inc. All rights reserved

EFFECT OF COVI-SOUP ON CD4 CELL COUNT AND HAEMOGLOBIN AMONG PATIENTS WITH DIABETES TYPE II AND HYPERTENSION IN KAKAMEGA COUNTY, KENYA: A RANDOMIZED CONTROL TRIAL

Type II diabetes mellitus (DM) and hypertension (HTN) diseases are part of the world’s challenges in achieving sustainable development goals. Cases of type II diabetes mellitus and hypertension diseases in low and middle-income countries are becoming a concern. Type II diabetes mellitus is an inflammatory disease. When there is inflammation the immune system is triggered and haemoglobin level is affected. While there have been numerous strategies to address the issue, these efforts have not been sufficient to guarantee a reduction in disease severity. This was an experimental study conducted in Kakamega County, Kenya. It employed a randomized control trial approach, with two groups: treatment and control. Systematic random sampling was used in identifying participants and simple randomization was used to allocate the respondent to the intervention groups. Sample sizes of 34 (treatment group) and 30 (control group) were used to produce quantitative data. This study evaluated the effect of COVI-soup on haemoglobin (Hb) and cluster of differentiation 4 (CD4) cell count of patients living with both type II diabetes mellitus and hypertension. The treatment group received COVI-soup (a soup composed of butternut squash, ginger, garlic, sunflower pepper, chia seeds and turmeric) while the control group was on a rice soup. Statistical Packages for Social Science (SPSS) version 20 was used in the data analysis. A paired student t-test was used to determine the effect of COVI-soup on Hb and CD4 cell count. There was a positive effect in CD4 cell count in the treatment group (P =0.001) whereas there was no significant effect in the control group (P =0.267). The mean change in CD4 cell count in the treatment group was 97 cells/ul and in the control 75 cells/ul. There was a positive effect (increase) on hemoglobin levels in the treatment group (p=0.0001) whereas there was no significant effect in the control group (p =0.126). The hemoglobin mean change in the treatment group was 3.9 g/dl and in the control 0.5 g/dl. Therefore, COVI-soup had a positive effect on hemoglobin and CD4 cell count levels among patients with both hypertension and type II diabetes mellitus

EFFECT OF COVI-SOUP ON CD4 CELL COUNT AND HAEMOGLOBIN AMONG PATIENTS WITH DIABETES TYPE II AND HYPERTENSION IN KAKAMEGA COUNTY, KENYA: A RANDOMIZED CONTROL TRIAL

Type II diabetes mellitus (DM) and hypertension (HTN) diseases are part of the world’s challenges in achieving sustainable development goals. Cases of type II diabetes mellitus and hypertension diseases in low and middle-income countries are becoming a concern. Type II diabetes mellitus is an inflammatory disease. When there is inflammation the immune system is triggered and haemoglobin level is affected. While there have been numerous strategies to address the issue, these efforts have not been sufficient to guarantee a reduction in disease severity. This was an experimental study conducted in Kakamega County, Kenya. It employed a randomized control trial approach, with two groups: treatment and control. Systematic random sampling was used in identifying participants and simple randomization was used to allocate the respondent to the intervention groups. Sample sizes of 34 (treatment group) and 30 (control group) were used to produce quantitative data. This study evaluated the effect of COVI-soup on haemoglobin (Hb) and cluster of differentiation 4 (CD4) cell count of patients living with both type II diabetes mellitus and hypertension. The treatment group received COVI-soup (a soup composed of butternut squash, ginger, garlic, sunflower pepper, chia seeds and turmeric) while the control group was on a rice soup. Statistical Packages for Social Science (SPSS) version 20 was used in the data analysis. A paired student t-test was used to determine the effect of COVI-soup on Hb and CD4 cell count. There was a positive effect in CD4 cell count in the treatment group (P =0.001) whereas there was no significant effect in the control group (P =0.267). The mean change in CD4 cell count in the treatment group was 97 cells/ul and in the control 75 cells/ul. There was a positive effect (increase) on hemoglobin levels in the treatment group (p=0.0001) whereas there was no significant effect in the control group (p =0.126). The hemoglobin mean change in the treatment group was 3.9 g/dl and in the control 0.5 g/dl. Therefore, COVI-soup had a positive effect on hemoglobin and CD4 cell count levels among patients with both hypertension and type II diabetes mellitus