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Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes.

ABSTRACT: Cell transfection is a technique wherein foreign genetic molecules are delivered into cells. To elucidate distinct responses during cell genetic modification, methods to achieve transfection at the single-cell level are of great value. Herein, we developed an automated micropipette-based quantitative microinjection technology that can deliver precise amounts of materials into cells. The developed microinjection system achieved precise single-cell microinjection by pre-patterning cells in an array and controlling the amount of substance delivered based on injection pressure and time. The precision of the proposed injection technique was examined by comparing the fluorescence intensities of fluorescent dye droplets with a standard concentration and water droplets with a known injection amount of the dye in oil. Injection of synthetic modified mRNA (modRNA) encoding green fluorescence proteins or a cocktail of plasmids encoding green and red fluorescence proteins into human foreskin fibroblast cells demonstrated that the resulting green fluorescence intensity or green/red fluorescence intensity ratio were well correlated with the amount of genetic material injected into the cells. Single-cell transfection via the developed microinjection technique will be of particular use in cases where cell transfection is challenging and genetically modified of selected cells are desired.

SUBMITTER: Chow YT

PROVIDER: S-EPMC4828701 | biostudies-other | 2016 Apr

REPOSITORIES: biostudies-other

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Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes.

Chow Yu Ting YT Chen Shuxun S Wang Ran R Liu Chichi C Kong Chi-Wing CW Li Ronald A RA Cheng Shuk Han SH Sun Dong D

Scientific reports 20160412

Cell transfection is a technique wherein foreign genetic molecules are delivered into cells. To elucidate distinct responses during cell genetic modification, methods to achieve transfection at the single-cell level are of great value. Herein, we developed an automated micropipette-based quantitative microinjection technology that can deliver precise amounts of materials into cells. The developed microinjection system achieved precise single-cell microinjection by pre-patterning cells in an arra ...[more]

PMID: 27067121

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Project description:Measuring the temperature of a sample is a fundamental need in many biological and chemical processes. When the volume of the sample is on the microliter or nanoliter scale (e.g., cells, microorganisms, precious samples, or samples in microfluidic devices), accurate measurement of the sample temperature becomes challenging. In this work, we demonstrate a technique for accurately determining the temperature of microliter volumes using a simple 3D-printed microfluidic chip. We accomplish this by first filling "microfluidic thermometer" channels on the chip with substances with precisely known freezing/melting points. We then use a thermoelectric cooler to create a stable and linear temperature gradient along these channels within a measurement region on the chip. A custom software tool (available as online Supporting Information) is then used to find the locations of solid-liquid interfaces in the thermometer channels; these locations have known temperatures equal to the freezing/melting points of the substances in the channels. The software then uses the locations of these interfaces to calculate the temperature at any desired point within the measurement region. Using this approach, the temperature of any microliter-scale on-chip sample can be measured with an uncertainty of about a quarter of a degree Celsius. As a proof-of-concept, we use this technique to measure the unknown freezing point of a 50 microliter volume of solution and demonstrate its feasibility on a 400 nanoliter sample. Additionally, this technique can be used to measure the temperature of any on-chip sample, not just near-zero-Celsius freezing points. We demonstrate this by using an oil that solidifies near room temperature (coconut oil) in a microfluidic thermometer to measure on-chip temperatures well above zero Celsius. By providing a low-cost and simple way to accurately measure temperatures in small volumes, this technique should find applications in both research and educational laboratories.

Dataset Information

Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes.

Publications

Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes.

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