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Deterministic bead-in-droplet ejection utilizing an integrated plug-in bead dispenser for single bead – based applications

View Article: PubMed Central - PubMed

ABSTRACT

This paper presents a deterministic bead-in-droplet ejection (BIDE) technique that regulates the precise distribution of microbeads in an ejected droplet. The deterministic BIDE was realized through the effective integration of a microfluidic single-particle handling technique with a liquid dispensing system. The integrated bead dispenser facilitates the transfer of the desired number of beads into a dispensing volume and the on-demand ejection of bead-encapsulated droplets. Single bead–encapsulated droplets were ejected every 3 s without any failure. Multiple-bead dispensing with deterministic control of the number of beads was demonstrated to emphasize the originality and quality of the proposed dispensing technique. The dispenser was mounted using a plug-socket type connection, and the dispensing process was completely automated using a programmed sequence without any microscopic observation. To demonstrate a potential application of the technique, bead-based streptavidin–biotin binding assay in an evaporating droplet was conducted using ultralow numbers of beads. The results evidenced the number of beads in the droplet crucially influences the reliability of the assay. Therefore, the proposed deterministic bead-in-droplet technology can be utilized to deliver desired beads onto a reaction site, particularly to reliably and efficiently enrich and detect target biomolecules.

No MeSH data available.


Trapping time variation at different bead concentrations.Red-solid, blue-solid, and black-empty bars indicate 600,000, 200,000, and 50,000 beads/mL, respectively.
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f4: Trapping time variation at different bead concentrations.Red-solid, blue-solid, and black-empty bars indicate 600,000, 200,000, and 50,000 beads/mL, respectively.

Mentions: Trapping time is defined as the time until the trapping of a following bead after the release of a trapped bead. Therefore, trapping time depends on the flux of the influent beads and is thus a function of the flow rate and bead concentration. We measured the trapping time at various bead concentrations at the optimized operation condition (i.e., at constant flow rate). To reduce operating time, we followed “trap–release–dispense–load” mode sequence rather than the “trap–release–load–dispense” sequence (Supplemental video clip 1). This change in the sequenced enabled faster reloading (by tens of milliseconds) of the following bead on the loading site; it also minimizes the time required for the entire process by enabling parallel operation of the modes: during continuous ejections, loading and trapping can occur concurrently. After a simple feasibility test, we ran the trapping test under the assumption that trapping a single bead requires less than 10 s. The time for each mode were set as follows: 10 s for trapping, 15 ms for releasing, and 30 ms for dispensing. The entire process was continually repeated for approximately 15 min (i.e., 90 droplet ejections). After 3 min, by which time the beads pipetted into the inlet reservoir reach the bead trap in the dispenser, the data for the trapping time was plotted (Fig. 4). As the bead concentration increased, the trapping time tended to decrease. At 50,000 beads/mL, the average trapping time was 6.5 s (standard deviation (SD) 10.22 s, maximum (MAX) 60 s, minimum (MIN) 4 s). Trapping failure occurred frequently (18%). The average trapping time was 1.05 s (SD 0.62 s, MAX 3.52 s, MIN 0.38 s) at 200,000 beads/mL and 0.71 s (SD 0.37 s, MAX 1.7 s, MIN 0.2 s) at 600,000 beads/mL. Although the trapping time decreased as bead concentration increased, a higher concentration occasionally resulted in channel clogging. Defect-free operation (i.e., 100% dispensing of droplets, each of which encapsulated a single bead) was realized when the trapping time was set sufficiently longer than the measured maximum trapping time for each concentration. At 600,000 beads/mL, we chose a trapping time of 3 s, considering the maximum trapping time of 1.7 s and an approximately 2-fold safety factor. Under the optimized operating conditions, our plug-in bead dispensing system was used for more than 300 ejections of droplets encapsulating a single bead, thereby confirming defect-free operation (Supplemental video clip 2).


Deterministic bead-in-droplet ejection utilizing an integrated plug-in bead dispenser for single bead – based applications
Trapping time variation at different bead concentrations.Red-solid, blue-solid, and black-empty bars indicate 600,000, 200,000, and 50,000 beads/mL, respectively.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5385560&req=5

f4: Trapping time variation at different bead concentrations.Red-solid, blue-solid, and black-empty bars indicate 600,000, 200,000, and 50,000 beads/mL, respectively.
Mentions: Trapping time is defined as the time until the trapping of a following bead after the release of a trapped bead. Therefore, trapping time depends on the flux of the influent beads and is thus a function of the flow rate and bead concentration. We measured the trapping time at various bead concentrations at the optimized operation condition (i.e., at constant flow rate). To reduce operating time, we followed “trap–release–dispense–load” mode sequence rather than the “trap–release–load–dispense” sequence (Supplemental video clip 1). This change in the sequenced enabled faster reloading (by tens of milliseconds) of the following bead on the loading site; it also minimizes the time required for the entire process by enabling parallel operation of the modes: during continuous ejections, loading and trapping can occur concurrently. After a simple feasibility test, we ran the trapping test under the assumption that trapping a single bead requires less than 10 s. The time for each mode were set as follows: 10 s for trapping, 15 ms for releasing, and 30 ms for dispensing. The entire process was continually repeated for approximately 15 min (i.e., 90 droplet ejections). After 3 min, by which time the beads pipetted into the inlet reservoir reach the bead trap in the dispenser, the data for the trapping time was plotted (Fig. 4). As the bead concentration increased, the trapping time tended to decrease. At 50,000 beads/mL, the average trapping time was 6.5 s (standard deviation (SD) 10.22 s, maximum (MAX) 60 s, minimum (MIN) 4 s). Trapping failure occurred frequently (18%). The average trapping time was 1.05 s (SD 0.62 s, MAX 3.52 s, MIN 0.38 s) at 200,000 beads/mL and 0.71 s (SD 0.37 s, MAX 1.7 s, MIN 0.2 s) at 600,000 beads/mL. Although the trapping time decreased as bead concentration increased, a higher concentration occasionally resulted in channel clogging. Defect-free operation (i.e., 100% dispensing of droplets, each of which encapsulated a single bead) was realized when the trapping time was set sufficiently longer than the measured maximum trapping time for each concentration. At 600,000 beads/mL, we chose a trapping time of 3 s, considering the maximum trapping time of 1.7 s and an approximately 2-fold safety factor. Under the optimized operating conditions, our plug-in bead dispensing system was used for more than 300 ejections of droplets encapsulating a single bead, thereby confirming defect-free operation (Supplemental video clip 2).

View Article: PubMed Central - PubMed

ABSTRACT

This paper presents a deterministic bead-in-droplet ejection (BIDE) technique that regulates the precise distribution of microbeads in an ejected droplet. The deterministic BIDE was realized through the effective integration of a microfluidic single-particle handling technique with a liquid dispensing system. The integrated bead dispenser facilitates the transfer of the desired number of beads into a dispensing volume and the on-demand ejection of bead-encapsulated droplets. Single bead–encapsulated droplets were ejected every 3 s without any failure. Multiple-bead dispensing with deterministic control of the number of beads was demonstrated to emphasize the originality and quality of the proposed dispensing technique. The dispenser was mounted using a plug-socket type connection, and the dispensing process was completely automated using a programmed sequence without any microscopic observation. To demonstrate a potential application of the technique, bead-based streptavidin–biotin binding assay in an evaporating droplet was conducted using ultralow numbers of beads. The results evidenced the number of beads in the droplet crucially influences the reliability of the assay. Therefore, the proposed deterministic bead-in-droplet technology can be utilized to deliver desired beads onto a reaction site, particularly to reliably and efficiently enrich and detect target biomolecules.

No MeSH data available.