How to do a SlipChip experiment:
The capability to create thousands of nanoliter-sized compartments is essential to isolating single molecules. (See associated video below)
Thank you to the 6-year-old volunteer and thank you to Liang Li at SlipChip Corp. for providing the chip used in this video.
Reference: Daan Witters, Bing Sun, Stefano Begolo, Jesus Rodriguez-Manzano, Whitney Robles, and Rustem F. Ismagilov, "Digital Biology and Chemistry," Lab on a Chip 2014, DOI: 10.1039/C4LC00248B.
Single-Molecule Counting on SlipChip Using a Mobile Phone:
In the video below, single molecules were amplified on SlipChip and a cell phone was used to image the chip in a shoebox. The image was then sent to a remote server, where the pattern of "positive" and "negative" wells on the chip was automatically analyzed, Poisson statistics was applied, and the number of molecules present in the sample was calculated. Quantitative results were automatically sent via email. A minimally trained user can use this approach.
Reference: David A. Selck, Mikhail A. Karymov, Bing Sun, and Rustem F. Ismagilov, "Increased Robustness of Single-Molecule Counting with Microfluidics, Digital Isothermal Amplification, and a Mobile Phone versus Real-Time Kinetic Measurements," Analytical Chemistry 2013, DOI: 10.1021/ac4030413.
Preloaded SlipChip:
In a preloaded SlipChip, the bottom plate contains wells preloaded with many reagents (different preloaded reagents are indicated by different shades of blue and green); in the assembled SlipChip these wells are covered by the top plate that acts as a lid for the wells with reagents. The device also contains a fluidic path composed of ducts in the bottom plate and wells in the top plate, which is connected only when the top and bottom plate are aligned in a specific configuration. Sample (white) can be injected into the fluidic path, filling both wells and ducts. Then, the top plate is "slipped", or moved, relative to the bottom plate, separating the sample into discrete volumes. The slipping step also allows complementary patterns of wells in both plates to overlap, exposing the sample-containing wells of the top plate to the reagent-containing wells of the bottom plate, and enables diffusion and reactions (indicated by a color change).
Reference: Wenbin Du, Liang Li, Kevin P. Nichols, and Rustem F. Ismagilov, "SlipChip", Lab Chip 2009 9: 2286-2292
i. single-step:
ii. multi-step:
Digital PCR SlipChip:
The digital PCR SlipChip is assembled such that elongated wells in the top (black) and bottom (red) plates overlap to form a continuous fluidic path. An aqueous reagent (violet) is injected into the SlipChip and fills the chip through the connected elongated wells. The top plate is slipped relative to the bottom plate such that the fluidic path is broken up and the circular wells are overlaid with the elongated wells, and aqueous droplets are formed in each compartment.
Reference: Feng Shen, Wenbin Du, Jason E. Kreutz, Alice Fok, and Rustem F. Ismagilov, "Digital PCR on a SlipChip", Lab Chip 2010 10: 2666-2672
Digital Isothermal Amplification:
The movie below shows schematic drawings of a two-step SlipChip for digital isothermal recombinase polymerase amplification (RPA). A first reaction mixture (red) is loaded, and slipping breaks the first fluidic path and compartmentalizes the loaded reagent. At the same time, the second fluidic path is formed by connecting type II wells, and the second reaction mixture (light blue) is loaded through a second inlet. An additional slipping step compartmentalizes reaction mixture 2 into the type II wells and overlaps the type II wells with the type I wells. The two reagents are mixed within the reaction compartments.
Reference: Feng Shen, Elena K. Davydova, Wenbin Du, Jason E. Kreutz, Olaf Piepenburg, and Rustem F. Ismagilov, "Digital Isothermal Quantification of Nucleic Acids via Simultaneous Chemical Initiation of Recombinase Polymerase Amplification Reactions on SlipChip", Analytical Chemistry 2011 83: 3533-3540
Filling and Spontaneous Flow:
Loading of a SlipChip via dead-end filling
In dead-end filling of a SlipChip, the lubricating fluid that fills the chip after assembly is dissipated through the gap between the two plates of
the SlipChip instead of flowing through an outlet at the end of the fluidic path. The movie below demonstrates dead-end filling of a SlipChip, where the green solution represents the sample, and the orange solution represents one reagent. The movie was generated from a series of consequential images taken with 1 s time intervals. The time duration of the whole loading process is close to 117 s.
Reference: Liang Li, Mikhail A. Karymov, Kevin P. Nichols, and Rustem F. Ismagilov, "Dead-End Filling of SlipChip Evaluated Theoretically and Experimentally as a Function of the Surface Chemistry and the Gap Size between the Plates for Lubricated and Dry SlipChips", Langmuir 2010 26: 12465-12471
Spontaneous flow
As demonstrated in the series of movies below, capillary pressure can be used to initiate and control the rate of spontaneous liquid-liquid flow through microfluidic channels.
Reference: Rebecca R. Pompano, Carol E. Platt, Mikhail A. Karymov, and Rustem F. Ismagilov, "Control of Initiation, Rate, and Routing of Spontaneous Capillary-Driven Flow of Liquid Droplets through Microfluidic Channels on SlipChip," Langmuir 2012 28: 1931-1941
i. Aqueous droplets flowing spontaneously on fast-flow chip
ii. Aqueous droplets flowing spontaneously on slow-flow chip
iii. Droplets of citrated human whole blood flowed spontaneously on a FEP-dip coated chip
User-loaded SlipChip:
In a user-loaded SlipChip, a sample and multiple reagents can be individually loaded through different fluidic paths that are created when the SlipChip is aligned in different configurations. Initially, the SlipChip is assembled so that the sample (purple) is loaded via one continuous fluidic path. The top plate is then moved, or "slipped" relative to the bottom plate, breaking up the sample into discrete volumes and connecting the fluidic paths for reagents. Different reagents (blue and brown) are loaded via separate fluidic paths. A second slipping step separates the reagents into discrete volumes and mixes the sample with different reagents. Different mixing ratios (determined by different well volumes) for each reagent are screened at the same time.
Reference: Liang Li, Wenbin Du, and Rustem F. Ismagilov,"User-Loaded SlipChip for Equipment-Free Multiplexed Nanoliter-Scale Experiments", JACS 2010 132: 106-111
FID SlipChip:
In the FID SlipChip, the sample and multiple reagents are individually loaded as in the user-loaded SlipChip. The movie shows slipping and diffusion in one set of wells containing one reagent with different equilibration times. After the sample (yellow) and reagent (blue) is loaded via a continuous fluidic path, the top plate (outlined in black) is slipped relative to the bottom plate (outlined in red). In this design, all wells are in the bottom plate. Upon slipping, the sample and reagents are separated into discrete volumes, and ducts in the top plate that were used to load the sample connect the sample wells to the reagent wells. Because the ducts are different lengths and widths, the equilibration time is varied across the SlipChip. The diffusion is shown in a time-lapse format during the second half of the movie. Different equilibration times are screened for each reagent at the same time: reagents (blue) diffuse quickly through shorter, wider ducts (left) and slowly through longer, narrower ducts (right).
Reference: Liang Li, Wenbin Du, and Rustem F. Ismagilov, "Multiparameter Screening on SlipChip Used for Nanoliter Protein Crystallization Combining Free Interface Diffusion and Microbatch Methods", JACS 2010 132: 112-119
SlipChip Device for Dry Sample Preservation in Remote Settings:
This SlipChip is used for dry preservation of biological specimens at room temperature.
Reference: Stefano Begolo, Feng Shen, and Rustem F. Ismagilov, "A Microfluidic Device for Dry Sample Preservation in Remote Settings," Lab Chip 2013 13: 4331-4342
Filling the sample preservation device
An untrained user can fill the dry sample preservation device. Once the user slips the device, the device can be shipped to a laboratory for analysis.
Dynamics of drying
The video below shows the process of sample drying within the device. Movie plays 60x faster than real-time.
Sample re-collection
The video below shows the steps for rehydrating and recovering dried samples. Rehydration and re-collection are shown only for one well, demonstrating the capability for partial recovery. Rehydration section plays 8x faster than real-time.