Design of an automated capillary electrophoresis platform for single-cell analysis

Abraham, David H.; Anttila, Matthew M.; Gallion, Luke; Petersen, Brae V.; Proctor, Angela; Allbritton, Nancy L.

doi:10.1016/bs.mie.2019.06.016

Cited by 8 publications

(11 citation statements)

References 62 publications

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“…Then, experimental throughput of the NCC technique for monocyte H 2 O 2 efflux monitoring is at an estimated~600 cells/h for single-channel operation with real-time single cell resolution. This appears to be at the highest range demonstrated to date for conventional chemical cytometry with reported values are 50-500 cells/h [27][28][29] . The previous reports also excluded labeling process time (usually taking a few hours) based on only the data acquisition time 24 .…”

Section: Resultsmentioning

confidence: 72%

“…Flow and chemical cytometry has been widely used to quantify the molecular heterogeneities of target cell populations. While typical flow and image cytometry of living cells can sample 10 6 -10 7 cells in just a few minutes [24][25][26] , the state of the art for the emerging field of chemical cytometry is between 50 and 500 cells/h since cells need to be pre-labeled, lysed, and separated to be detected [27][28][29] . Nevertheless, this level of throughput has elevated chemical cytometry as a valuable cell characterization tool allowing quantitative information to be gathered with high selectivity and signal-to-noise ratio 30,31 .…”

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confidence: 99%

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Cellular lensing and near infrared fluorescent nanosensor arrays to enable chemical efflux cytometry

et al. 2021

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Nanosensors have proven to be powerful tools to monitor single cells, achieving spatiotemporal precision even at molecular level. However, there has not been way of extending this approach to statistically relevant numbers of living cells. Herein, we design and fabricate nanosensor array in microfluidics that addresses this limitation, creating a Nanosensor Chemical Cytometry (NCC). nIR fluorescent carbon nanotube array is integrated along microfluidic channel through which flowing cells is guided. We can utilize the flowing cell itself as highly informative Gaussian lenses projecting nIR profiles and extract rich information. This unique biophotonic waveguide allows for quantified cross-correlation of biomolecular information with various physical properties and creates label-free chemical cytometer for cellular heterogeneity measurement. As an example, the NCC can profile the immune heterogeneities of human monocyte populations at attomolar sensitivity in completely non-destructive and real-time manner with rate of ~600 cells/hr, highest range demonstrated to date for state-of-the-art chemical cytometry.

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Section: Resultsmentioning

confidence: 72%

mentioning

confidence: 99%

Cellular lensing and near infrared fluorescent nanosensor arrays to enable chemical efflux cytometry

et al. 2021

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“…This appears to be superior to range demonstrated to date for conventional chemical cytometry where reported values are 50 -500 cells/hr. [27][28][29] Our next steps for NCC will utilize a parallel and multiple channel approach, automated fluidic control coupled to automated data collection to increase this throughput substantially. Preliminary work showed that analyzing cell number could be improved with 300% increasement by using three parallel channels of a single microfluidic chip ( Figure S25).…”

Section: Ncc For Monitoring Of Multimodal Immune Response Heterogeneimentioning

confidence: 99%

“…While typical flow and image cytometry of living cells can sample 10 6 -10 7 cells in just a few minutes, [24][25][26] the state of the art for the emerging field of chemical cytometry is between 50 to 500 cells/hr since cells need to be pre-labelled, lysed, and separated to be detected. [27][28][29] Nevertheless, this level of throughput has elevated chemical cytometry as a valuable cell characterization tool allowing quantitative information to be gathered with high selectivity and signal-to-noise ratio. 30,31 Nanosensors have significant potential to greatly expand the number of variables measured in chemical cytometry given the large number of new types being demonstrated in the recent literature.…”

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confidence: 99%

Cellular Lensing and Near Infrared Fluorescent Nanosensor Arrays to Enable Chemical Efflux Cytometry

Cho

Gong

Koman

et al. 2020

Preprint

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Nanosensor have proven to be powerful tools to monitor single biological cells and organisms, achieving spatial and temporal precision even at the single molecule level. However, there has not been a way of extending this approach to statistically relevant numbers of living cells and organisms. Herein, we design and fabricate a high throughput nanosensor array in a microfluidic channel that addresses this limitation, creating a Nanosensor Chemical Cytometry (NCC). An array of nIR fluorescent single walled carbon nanotube (SWNT) nanosensors is integrated along a microfluidic channel through which a population of flowing cells is guided. We show that one can utilize the flowing cell itself as highly informative Gaussian lenses projecting nIR emission profiles and extract rich information on a per cell basis at high throughput. This unique biophotonic waveguide allows for quantified cross-correlation of the biomolecular information with physical properties such as cellular diameter, refractive index (RI), and eccentricity and creates a label-free chemical cytometer for the measurement of cellular heterogeneity with unprecedented precision. As an example, the NCC can profile the immune response heterogeneities of distinct human monocyte populations at attomolar (10-18 moles) sensitivity in a completely non-destructive and real-time manner with a rate of ~100 cells/frame, highest range demonstrated to date for state of the art chemical cytometry. We demonstrate distinct H2O2 efflux heterogeneities between 330 and 624 attomole/cell·min with cell projected areas between 271 and 263 µm2, eccentricity values between 0.405 and 0.363 and RI values between 1.383 and 1.377 for non-activated and activated human monocytes, respectively. Hence, we show that our nanotechnology based biophotonic cytometer has significant potential and versatility to answer important questions and provide new insight in immunology, cell manufacturing and biopharmaceutical research.

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“…Several fluorescent detection setups have been described for chemical cytometry, many of which have employed photomultiplier tubes (PMTs). [9][10][11] PMTs work using a photocathode housed within a vacuum environment to transduce light into an electrical signal. PMTs have high gain (10 5-9 ) and maintain linearity over a large dynamic range, typically as many as 6 orders of magnitude.…”

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confidence: 99%

Silicon Photomultipliers as a Low-Cost Fluorescence Detector for Capillary Electrophoresis

Petersen¹,

Gallion²,

Allbritton³

2020

Preprint

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Capillary electrophoresis (CE) is a highly efficient separation method capable of handling small sample volumes (~pL) and low (~yoctomole) detection limits, and as such is ideal for applications that require high sensitivity such as single-cell analysis. Low-cost CE instrumentation is quickly expanding but low-cost, open-source fluorescence detectors with ultra-sensitive detection limits are lacking. Silicon photomultipliers (SiPM) are inexpensive, low-footprint detectors with the potential to fill the role as a detector when cost, size, and customization are important. In this work we demonstrate the use of a SiPM in CE with zeptomolar detection limits and a dynamic range spanning five orders of magnitude, comparable to photomultiplier detectors. We characterize the performance of the SiPM as a highly sensitive detector by measuring enzyme activity in single cells. This simple, small footprint, and low-cost (<$130) light detection circuit will be beneficial for open-source, portable, and budget friendly instrumentation requiring high sensitivity.<br>

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Design of an automated capillary electrophoresis platform for single-cell analysis

Cited by 8 publications

References 62 publications

Cellular lensing and near infrared fluorescent nanosensor arrays to enable chemical efflux cytometry

Cellular lensing and near infrared fluorescent nanosensor arrays to enable chemical efflux cytometry

Cellular Lensing and Near Infrared Fluorescent Nanosensor Arrays to Enable Chemical Efflux Cytometry

Silicon Photomultipliers as a Low-Cost Fluorescence Detector for Capillary Electrophoresis

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