@article {200, title = {Electricity-free amplification and detection for molecular point-of-care diagnosis of HIV-1.}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014}, pages = {e113693}, abstract = {

In resource-limited settings, the lack of decentralized molecular diagnostic testing and sparse access to centralized medical facilities can present a critical barrier to timely diagnosis, treatment, and subsequent control and elimination of infectious diseases. Isothermal nucleic acid amplification methods, including reverse transcription loop-mediated isothermal amplification (RT-LAMP), are well-suited for decentralized point-of-care molecular testing in minimal infrastructure laboratories since they significantly reduce the complexity of equipment and power requirements. Despite reduced complexity, however, there is still a need for a constant heat source to enable isothermal nucleic acid amplification. This requirement poses significant challenges for laboratories in developing countries where electricity is often unreliable or unavailable. To address this need, we previously developed a low-cost, electricity-free heater using an exothermic reaction thermally coupled with a phase change material. This heater achieved acceptable performance, but exhibited considerable variability. Furthermore, as an enabling technology, the heater was an incomplete diagnostic solution. Here we describe a more precise, affordable, and robust heater design with thermal standard deviation <0.5{\textdegree}C at operating temperature, a cost of approximately US$.06 per test for heater reaction materials, and an ambient temperature operating range from 16{\textdegree}C to 30{\textdegree}C. We also pair the heater with nucleic acid lateral flow (NALF)-detection for a visual readout. To further illustrate the utility of the electricity-free heater and NALF-detection platform, we demonstrate sensitive and repeatable detection of HIV-1 with a {\ss}-actin positive internal amplification control from processed sample to result in less than 80 minutes. Together, these elements are building blocks for an electricity-free platform capable of isothermal amplification and detection of a variety of pathogens.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0113693}, author = {Singleton, Jered and Osborn, Jennifer L and Lillis, Lorraine and Hawkins, Kenneth and Guelig, Dylan and Price, Will and Johns, Rachel and Ebels, Kelly and Boyle, David and Weigl, Bernhard and LaBarre, Paul} } @article {207, title = {Isothermal amplification using a chemical heating device for point-of-care detection of HIV-1.}, journal = {PLoS One}, volume = {7}, year = {2012}, month = {2012}, pages = {e31432}, abstract = {

BACKGROUND: To date, the use of traditional nucleic acid amplification tests (NAAT) for detection of HIV-1 DNA or RNA has been restricted to laboratory settings due to time, equipment, and technical expertise requirements. The availability of a rapid NAAT with applicability for resource-limited or point-of-care (POC) settings would fill a great need in HIV diagnostics, allowing for timely diagnosis or confirmation of infection status, as well as facilitating the diagnosis of acute infection, screening and evaluation of infants born to HIV-infected mothers. Isothermal amplification methods, such as reverse-transcription, loop-mediated isothermal amplification (RT-LAMP), exhibit characteristics that are ideal for POC settings, since they are typically quicker, easier to perform, and allow for integration into low-tech, portable heating devices.

METHODOLOGY/SIGNIFICANT FINDINGS: In this study, we evaluated the HIV-1 RT-LAMP assay using portable, non-instrumented nucleic acid amplification (NINA) heating devices that generate heat from the exothermic reaction of calcium oxide and water. The NINA heating devices exhibited stable temperatures throughout the amplification reaction and consistent amplification results between three separate devices and a thermalcycler. The performance of the NINA heaters was validated using whole blood specimens from HIV-1 infected patients.

CONCLUSION: The RT-LAMP isothermal amplification method used in conjunction with a chemical heating device provides a portable, rapid and robust NAAT platform that has the potential to facilitate HIV-1 testing in resource-limited settings and POC.

}, keywords = {AIDS Serodiagnosis, DNA Primers, DNA, Viral, Equipment Design, Heating, HIV Infections, HIV-1, Humans, Nucleic Acid Amplification Techniques, Point-of-Care Systems, RNA, Viral, Temperature}, issn = {1932-6203}, doi = {10.1371/journal.pone.0031432}, author = {Curtis, Kelly A and Rudolph, Donna L and Nejad, Irene and Singleton, Jered and Beddoe, Andy and Weigl, Bernhard and LaBarre, Paul and Owen, S Michele} } @patent {marsot_united_2012, title = {United {States} {Patent}: 8282576 - {Method} and apparatus for an improved sample capture device}, number = {8282576}, year = {2012}, abstract = {A body fluid sampling device is provided. A mesh may be used to guide blood or fluid to travel directly from the wound to an analyte detecting port on the cartridge. Thus the volume of blood or body fluid produced at the wound site irregardless of its droplet geometry can be reliable and substantially completely transported to the analyte detecting member for measurement.}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&p=1\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-bool.html\&r=11\&f=G\&l=50\&co1=AND\&d=PTXT\&s1=\%22Bardell\%3B+Ron\%22.INNM.\&OS=IN/\%22Bardell;+Ron\%22\&RS=IN/\%22Bardell;+Ron\%22}, author = {Marsot, Travis and Lum, Paul and Alden, Don and Ross, James and Bardell, Ron L. and Weigl, Bernhard} } @article {208, title = {Gestational diabetes screening: the low-cost algorithm.}, journal = {Int J Gynaecol Obstet}, volume = {115 Suppl 1}, year = {2011}, month = {2011 Nov}, pages = {S30-3}, abstract = {

The American Diabetes Association has endorsed the demanding recommendation by the International Association of the Diabetes and Pregnancy Study Groups (IADPSG) that every pregnant woman should undergo the oral glucose tolerance test (OGTT) for the screening of gestational diabetes mellitus (GDM). The aim of this study was to find out if the fasting plasma glucose (FPG) and newer emerging technologies could simplify the cumbersome IADPSG algorithm. Two FPG thresholds (of the OGTT) were used to rule in and rule out GDM in the Hyperglycemia and Adverse Pregnancy Outcome (HAPO) cohort (n = 23316) and a population at high risk for GDM (n = 10283). For the HAPO cohort and the high-risk population, respectively, FPG thresholds of: (a) >= 5.1 mmol/L (specificity 100\%) independently ruled in GDM in 1769 (8.3\%) women and 2975 (28.9\%) women; and (b) <= 4.4 mmol/L ruled out GDM in 11526 (49.4\%) women (84.1\% sensitivity) and 2228 (21.7\%) women (95.4\% sensitivity). Use of the FPG independently could have avoided 13295 (57.0\%) and 5203 (50.6\%) OGTTs in the 2 groups. The initial FPG-by significantly reducing the number of cumbersome OGTTs needed-can make the IADPSG recommendations more acceptable worldwide. The number of GDM women missed is population dependent. For low-resource countries, alternative newer and cheaper tests in development hold an exciting future.

}, keywords = {Algorithms, Diabetes, Gestational, Fasting, Female, Glucose Tolerance Test, Guidelines as Topic, Humans, Hyperglycemia, Mass Screening, Pregnancy, Pregnancy Complications, Sensitivity and Specificity}, issn = {1879-3479}, doi = {10.1016/S0020-7292(11)60009-X}, author = {Agarwal, Mukesh M and Weigl, Bernhard and Hod, Moshe} } @article {210, title = {Instrument-free nucleic acid amplification assays for global health settings.}, journal = {Proc SPIE Int Soc Opt Eng}, volume = {8029}, year = {2011}, month = {2011 May 16}, abstract = {

Many infectious diseases that affect global health are most accurately diagnosed through nucleic acid amplification and detection. However, existing nucleic acid amplification tests are too expensive and complex for most low-resource settings. The small numbers of centralized laboratories that exist in developing countries tend to be in urban areas and primarily cater to the affluent. In contrast, rural area health care facilities commonly have only basic equipment and health workers have limited training and little ability to maintain equipment and handle reagents.(1) Reliable electric power is a common infrastructure shortfall. In this paper, we discuss a practical approach to the design and development of non-instrumented molecular diagnostic tests that exploit the benefits of isothermal amplification strategies. We identify modular instrument-free technologies for sample collection, sample preparation, amplification, heating, and detection. By appropriately selecting and integrating these instrument-free modules, we envision development of an easy to use, infrastructure independent diagnostic test that will enable increased use of highly accurate molecular diagnostics at the point of care in low-resource settings.

}, issn = {0277-786X}, doi = {10.1117/12.882868}, author = {LaBarre, Paul and Boyle, David and Hawkins, Kenneth and Weigl, Bernhard} } @article {209, title = {A simple, inexpensive device for nucleic acid amplification without electricity-toward instrument-free molecular diagnostics in low-resource settings.}, journal = {PLoS One}, volume = {6}, year = {2011}, month = {2011}, pages = {e19738}, abstract = {

BACKGROUND: Molecular assays targeted to nucleic acid (NA) markers are becoming increasingly important to medical diagnostics. However, these are typically confined to wealthy, developed countries; or, to the national reference laboratories of developing-world countries. There are many infectious diseases that are endemic in low-resource settings (LRS) where the lack of simple, instrument-free, NA diagnostic tests is a critical barrier to timely treatment. One of the primary barriers to the practicality and availability of NA assays in LRS has been the complexity and power requirements of polymerase chain reaction (PCR) instrumentation (another is sample preparation).

METHODOLOGY/PRINCIPAL FINDINGS: In this article, we investigate the hypothesis that an electricity-free heater based on exothermic chemical reactions and engineered phase change materials can successfully incubate isothermal NA amplification assays. We assess the heater{\textquoteright}s equivalence to commercially available PCR instruments through the characterization of the temperature profiles produced, and a minimal method comparison. Versions of the prototype for several different isothermal techniques are presented.

CONCLUSIONS/SIGNIFICANCE: We demonstrate that an electricity-free heater based on exothermic chemical reactions and engineered phase change materials can successfully incubate isothermal NA amplification assays, and that the results of those assays are not significantly different from ones incubated in parallel in commercially available PCR instruments. These results clearly suggest the potential of the non-instrumented nucleic acid amplification (NINA) heater for molecular diagnostics in LRS. When combined with other innovations in development that eliminate power requirements for sample preparation, cold reagent storage, and readout, the NINA heater will comprise part of a kit that should enable electricity-free NA testing for many important analytes.

}, keywords = {Developing Countries, Electricity, Hot Temperature, Molecular Diagnostic Techniques, Nucleic Acid Amplification Techniques, Plasmodium falciparum, Reference Standards}, issn = {1932-6203}, doi = {10.1371/journal.pone.0019738}, author = {LaBarre, Paul and Hawkins, Kenneth R and Gerlach, Jay and Wilmoth, Jared and Beddoe, Andrew and Singleton, Jered and Boyle, David and Weigl, Bernhard} } @article {215, title = {Non-instrumented nucleic acid amplification (NINA): instrument-free molecular malaria diagnostics for low-resource settings.}, journal = {Conf Proc IEEE Eng Med Biol Soc}, volume = {2010}, year = {2010}, month = {2010}, pages = {1097-9}, abstract = {

We have achieved the first complete, non-instrumented nucleic acid amplification test (NAAT) using a calcium oxide heat source thermally linked to an engineered phase change material. These two components alone maintain a thermal profile suitable for the loop-mediated isothermal amplification assay. Starting with computational fluid dynamics analysis, we identified nominal geometry for the exothermic reaction chamber, phase change material chamber, thermal insulation, and packaging. Using this model, we designed and fabricated an alpha prototype assay platform. We have verified the function of this multi-pathogen-capable platform with both fluorescent and visual turbidity indications using samples spiked with malaria DNA. Both the exothermically heated platform samples and samples heated on a Perkin-Elmer GeneAmp9600 thermocycler were first incubated at 62{\textdegree}C for 45 minutes, then heated to 95{\textdegree}C to terminate enzyme activity, then analyzed. Results from the exothermically heated, non-instrumented platform were comparable to those from the thermocycler. These developments will enable point-of-care diagnostics using accurate NAATs which until now have required a well-equipped laboratory. The aim of this research is to provide pathogen detection with NAAT-level sensitivity in low-resource settings where assays such as immunochromatographic strip tests are successfully used but where there is no access to the infrastructure and logistics required to operate and maintain instrument-based diagnostics.

}, keywords = {DNA, Protozoan, Heating, Humans, Malaria, Falciparum, Molecular Diagnostic Techniques, Nucleic Acid Amplification Techniques}, issn = {1557-170X}, doi = {10.1109/IEMBS.2010.5627346}, author = {LaBarre, Paul and Gerlach, Jay and Wilmoth, Jared and Beddoe, Andrew and Singleton, Jered and Weigl, Bernhard} } @article {219, title = {A new HPV-DNA test for cervical-cancer screening in developing regions: a cross-sectional study of clinical accuracy in rural China.}, journal = {Lancet Oncol}, volume = {9}, year = {2008}, month = {2008 Oct}, pages = {929-36}, abstract = {

BACKGROUND: A new test (careHPV; QIAGEN, Gaithersburg, MD, USA) has been developed to detect 14 high-risk types of carcinogenic human papillomavirus (HPV) in about 2.5 h, to screen women in developing regions for cervical intraepithelial neoplasia (CIN). We did a cross-sectional study to assess the clinical accuracy of careHPV as a rapid screening test in two county hospitals in rural China.

METHODS: From May 10 to June 15, 2007, the careHPV test was done locally by use of self-obtained vaginal and provider-obtained cervical specimens from a screening population-based set of 2530 women aged 30 to 54 years in Shanxi province, China. All women were assessed by visual inspection with acetic acid (VIA), Digene High-Risk HPV HC2 DNA Test (HC2), liquid-based cytology, and colposcopy with directed biopsy and endocervical curettage as necessary. In 2388 women with complete data, 441 women with negative colposcopy, but unsatisfactory or abnormal cytology or who were positive on HC2 or the new careHPV test, were recalled for a second colposcopy, four-quadrant cervical biopsies, and endocervical curettage. An absence of independence between the tests was not adjusted for and the Bonferroni correction was used for multiple comparisons.

FINDINGS: Complete data were available for 2388 (94.4\%) women. 70 women had CIN2+ (moderate or severe CIN or cancer), of whom 23 had CIN3+. By use of CIN2+ as the reference standard and area-under-the-curve analysis with a two-sided alpha error level of 0.0083, the sensitivities and specificities of the careHPV test for a cut-off ratio cut-point of 0.5 relative light units, were 90.0\% (95\% CI 83.0-97.0) and 84.2\% (82.7-85.7), respectively, on cervical specimens, and 81.4\% (72.3-90.5) and 82.4\% (80.8-83.9), respectively, on vaginal specimens (areas under the curve not significantly different, p=0.0596), compared with 41.4\% (29.9-53.0) and 94.5\% (93.6-95.4) for VIA (areas under the curve significantly different, p=0.0001 and p=0.0031, for cervical and vaginal-specimen comparisons for the careHPV test, respectively). The sensitivity and specificity of HC2 for cervical specimens were 97.1\% (93.2-100) and 85.6\% (84.2-87.1), respectively (areas under the curve not significantly different from the careHPV test on cervical specimens, p=0.0163).

INTERPRETATION: The careHPV test is promising as a primary screening method for cervical-cancer prevention in low-resource regions.

}, keywords = {Adult, Alphapapillomavirus, Cervical Intraepithelial Neoplasia, China, Cross-Sectional Studies, DNA, Viral, Female, Humans, Mass Screening, Middle Aged, Papillomavirus Infections, Predictive Value of Tests, ROC Curve, Rural Health Services, Sensitivity and Specificity, Tumor Markers, Biological, Uterine Cervical Neoplasms}, issn = {1474-5488}, doi = {10.1016/S1470-2045(08)70210-9}, author = {Qiao, You-Lin and Sellors, John W and Eder, Paul S and Bao, Yan-Ping and Lim, Jeanette M and Zhao, Fang-Hui and Weigl, Bernhard and Zhang, Wen-Hua and Peck, Roger B and Li, Ling and Chen, Feng and Pan, Qing-Jing and Lorincz, Attila T} } @article {220, title = {The NIBIB Point of Care Technologies Research Network Center Themes and Opportunities for Exploratory POC Projects.}, journal = {Point Care}, volume = {7}, year = {2008}, month = {2008 Mar}, pages = {1-41}, abstract = {

This article describes the new National Institute of Biomedical Imaging and Bioengineering Point-of-Care (POC) Technologies Research Network and its 4 Centers. The goal is to build expertise in development of integrated systems that address unmet POC testing clinical needs. Centers will work individually and also collectively as part of the national network to coordinate development, clinical evaluation, and reduction to practice of new POC devices.

}, issn = {1533-029X}, doi = {10.1097/POC.0b013e318162f3dd}, author = {Kost, Gerald J and Korte, Brenda and Beyette, Fred R and Gaydos, Charlotte and Weigl, Bernhard} } @article {218, title = {Towards non- and minimally instrumented, microfluidics-based diagnostic devices.}, journal = {Lab Chip}, volume = {8}, year = {2008}, month = {2008 Dec}, pages = {1999-2014}, abstract = {

In many health care settings, it is uneconomical, impractical, or unaffordable to maintain and access a fully equipped diagnostics laboratory. Examples include home health care, developing-country health care, and emergency situations in which first responders are dealing with pandemics or biowarfare agent release. In those settings, fully disposable diagnostic devices that require no instrument support, reagent, or significant training are well suited. Although the only such technology to have found widespread adoption so far is the immunochromatographic rapid assay strip test, microfluidics holds promise to expand the range of assay technologies that can be performed in formats similar to that of a strip test. In this paper, we review progress toward development of disposable, low-cost, easy-to-use microfluidics-based diagnostics that require no instrument at all. We also present examples of microfluidic functional elements--including mixers, separators, and detectors--as well as complete microfluidic devices that function entirely without any moving parts and external power sources.

}, keywords = {Diagnostic Techniques and Procedures, Disposable Equipment, Microfluidics}, issn = {1473-0197}, doi = {10.1039/b811314a}, author = {Weigl, Bernhard and Domingo, Gonzalo and LaBarre, Paul and Gerlach, Jay} } @patent {weigl_united_2007, title = {United {States} {Patent}: 7196842 - {Attenuating} filter for ultraviolet light}, number = {7196842}, year = {2007}, abstract = {An attenuating filter provides a prescribed attenuation of the intensity of transmitted, short-wavelength, ultraviolet light, in particular, at wavelengths below 200 nm, that is governed by a predefinable spatial distribution of its spectral transmittance. The filter has a transparent substrate (3), e.g. fabricated from crystalline calcium fluoride. A filter coating (5) fabricated from a dielectric material that absorbs over a predefined wavelength range is applied to at least one surface (4) of the substrate. In the case of operating wavelengths of about 193 nm, the filter coating consists largely of tantalum pentoxide. Filters of the type, which may be inexpensively fabricated with high yields, are noted for their high abilities to withstand laser radiation and may be effectively antireflection coated employing simply designed antireflection coatings.}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&p=1\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-bool.html\&r=16\&f=G\&l=50\&co1=AND\&d=PTXT\&s1=\%22Weigl\%3B+Bernhard\%22.INNM.\&OS=IN/\%22Weigl;+Bernhard\%22\&RS=IN/\%22Weigl;+Bernhard\%22}, author = {Weigl, Bernhard and DE and Paul, Hans-Jochen and DE and Eva, Eric and DE} } @patent {wu_united_2001, title = {United {States} {Patent}: 6221677 - {Simultaneous} particle separation and chemical reaction}, number = {6221677}, year = {2001}, abstract = {This invention provides a method and apparatus for detecting the presence of analyte particles in a sample fluid also comprising larger particles, particularly blood. It exploits diffusion to provide simultaneous filtering of the larger particles and reaction of the analyte particles. A sample stream and a reagent stream join on the upstream end of a laminar flow reaction channel and flow in adjacent laminar streams. The reagents can be in solution or immobilized on a bead. The analyte particles diffuse from the sample stream into the reagent stream, leaving behind the larger particles in the residual sample stream. In the reagent stream the analyte particles react with reagent particles and form product particles, thereby creating a product stream. At the downstream end of the reaction channel, the residual sample stream and the product stream are divided. The product particles are then detected, preferably optically, in the product stream. Prior to detection, the product stream can undergo further filtering or separation, or can join a second reagent stream to form secondary product particles. This apparatus and method can be used to implement competitive immunoassays or sandwich immunoassays using enzymatically or fluorescently labeled immunoreagents. The apparatus and method can also be used to synthesize products, in which case two reagent streams join in the laminar flow reaction channel.}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&p=1\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-bool.html\&r=29\&f=G\&l=50\&co1=AND\&d=PTXT\&s1=\%22Weigl\%3B+Bernhard\%22.INNM.\&OS=IN/\%22Weigl;+Bernhard\%22\&RS=IN/\%22Weigl;+Bernhard\%22}, author = {Wu, Caicai and Weigl, Bernhard and Kenny, Margaret A. and Yager, Paul} } @patent {weigl_united_2001, title = {United {States} {Patent}: 6294296 - {Method} and device for mutually aligning a mask pattern formed in a mask and a substrate}, number = {6294296}, year = {2001}, month = {sep}, abstract = {In a method for mutually aligning a mask pattern formed in a mask 1 and a substrate 2, on which the mask pattern is to be imaged, by using setting marks 12a, 12b and 13a or 13b in the mask 1 and in the substrate 2, the alignment is performed with the aid of an imaging system and a light beam with polarized light 9. A phase shift for the first diffraction orders 20 is undertaken in the beam path 9a, 9b. Higher diffraction orders 21 and unwanted light are filtered out after the phase shift, and after the filtering out the light beams of the first diffraction orders 20 are detected, and the result is evaluated for the purpose of alignment.}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&p=1\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-bool.html\&r=28\&f=G\&l=50\&co1=AND\&d=PTXT\&s1=\%22Weigl\%3B+Bernhard\%22.INNM.\&OS=IN/\%22Weigl;+Bernhard\%22\&RS=IN/\%22Weigl;+Bernhard\%22}, author = {Weigl, Bernhard and DE} } @patent {wu_united_2001-1, title = {United {States} {Patent}: 6297061 - {Simultaneous} particle separation and chemical reaction}, number = {6297061}, year = {2001}, month = {oct}, abstract = {This invention provides a method and apparatus for detecting the presence of analyte particles in a sample fluid also comprising larger particles, particularly blood. It exploits diffusion to provide simultaneous filtering of the larger particles and reaction of the analyte particles. A sample stream and a reagent stream join on the upstream end of a laminar flow reaction channel and flow in adjacent laminar streams. The reagents can be in solution or immobilized on a bead. The analyte particles diffuse from the sample stream into the reagent stream, leaving behind the larger particles in the residual sample stream. In the reagent stream the analyte particles react with reagent particles and form product particles, thereby creating a product stream. At the downstream end of the reaction channel, the residual sample stream and the product stream are divided. The product particles are then detected, preferably optically, in the product stream. Prior to detection, the product stream can undergo further filtering or separation, or can join a second reagent stream to form secondary product particles. This apparatus and method can be used to implement competitive immunoassays or sandwich immunoassays using enzymatically or fluorescently labeled immunoreagents. The apparatus and method can also be used to synthesize products, in which case two reagent streams join in the laminar flow reaction channel.}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&p=1\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-bool.html\&r=27\&f=G\&l=50\&co1=AND\&d=PTXT\&s1=\%22Weigl\%3B+Bernhard\%22.INNM.\&OS=IN/\%22Weigl;+Bernhard\%22\&RS=IN/\%22Weigl;+Bernhard\%22}, author = {Wu, Caicai and Weigl, Bernhard and Kenny, Margaret A. and Yager, Paul} } @patent {weigl_united_2000-2, title = {United {States} {Patent}: 6136272 - {Device} for rapidly joining and splitting fluid layers}, number = {6136272}, year = {2000}, month = {oct}, abstract = {A device and method for introducing a second laminar fluid layer to, or removing a second laminar fluid layer from, a first laminar fluid layer are provided. Each laminar fluid layer can contain two or more side by side laminar streams. The device includes a main flow channel, and at least one tributary channel in fluid connection with a bridge channel which is in fluid connection with main flow channel. The device can be formed in a single piece of material, which can be optically transparent. Optionally, the channels can be formed in a first plate, the first and optionally the second surfaces of which are sealed to a second and optionally a third plate. The second and third plates can be optically transparent to allow for optical detection and analysis. A first laminar fluid layer is introduced into the main flow channel. If a second laminar fluid layer is to be added to the first laminar fluid layer, then the former is introduced into the tributary channel, from whence it flows into the bridge channel and then into the main flow channel, where it flows below the first laminar fluid layer and diffusionally mixes with it. Preferably, the width of the main flow channel is relatively small, so that particles in an added second laminar fluid layer diffusionally mix into the first laminar fluid layer rapidly. If a second laminar fluid layer is to be removed from a first laminar fluid layer, then the latter is split into two portions: one portion continues flowing down the main flow channel and one portion flows into the bridge channel from whence it flows into the tributary channel.}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&p=1\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-bool.html\&r=32\&f=G\&l=50\&co1=AND\&d=PTXT\&s1=\%22Weigl\%3B+Bernhard\%22.INNM.\&OS=IN/\%22Weigl;+Bernhard\%22\&RS=IN/\%22Weigl;+Bernhard\%22}, author = {Weigl, Bernhard and Zebert, Diane M. and Kenny, Margaret A.} } @patent {weigl_united_2000-1, title = {United {States} {Patent}: 6159739 - {Device} and method for 3-dimensional alignment of particles in microfabricated flow channels}, number = {6159739}, year = {2000}, month = {dec}, abstract = {The present invention provides a sheath flow module made from a first plate of material having formed therein a laminar fluid flow channel; at least two inlets, each inlet joining the laminar flow channel at a junction, the first inlet junction being wider than the second inlet junction, and an outlet from the flow channel. A second plate, e.g. a transparent cover plate, seals the module and allows for optical measurements. A first inlet allows for introduction of a first fluid into the flow channel. The first fluid is the sheath fluid. A second inlet allows for introduction of a second fluid into the sheath fluid while it is flowing through the flow channel. The second fluid is the center fluid. Because the second inlet junction is narrower than the first inlet junction, the center fluid becomes surrounded on both sides by the sheath fluid. After all fluids have been introduced and sheath flow has been achieved, the depth of the flow channel can be decreased, leading to vertical hydrodynamic focusing. Optionally, the width of the flow channel can be decreased, leading to horizontal hydrodynamic focusing. The decrease in depth and width can be gradual or abrupt. The device of the present invention can function in two modes, the sheath flow mode and the particle injector mode, depending on the relative densities of the sheath fluid, the center fluid, and any particles in either fluid.}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&p=1\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-bool.html\&r=31\&f=G\&l=50\&co1=AND\&d=PTXT\&s1=\%22Weigl\%3B+Bernhard\%22.INNM.\&OS=IN/\%22Weigl;+Bernhard\%22\&RS=IN/\%22Weigl;+Bernhard\%22}, author = {Weigl, Bernhard and Yager, Paul and Brody, James P.} }