Sample reader with oil circulation

Abstract

Systems, including methods and devices, for analyzing partitioned samples, such as droplets, using recycled fluid. The system can be used to analyze samples of multiple zones, where fluid for an initial sample can be reused for subsequent samples. This reuse or recycle can reduce the amount of fluid required to perform the analysis, while reducing costs. Furthermore, in some embodiments, it may simplify operations by increasing the number of analyses that may be performed before the fluid has to be replenished or replaced.

Description

Cross-reference to priority application

[0001] This application is based upon and claims the benefit of 35 U.S.C. §119(e) U.S. Provisional Patent Application Serial No. 63 / 358,509, filed on July 5, 2022, which is incorporated herein by reference in its entirety for all purposes. introduction

[0002] Many biomedical applications rely on high-throughput determination of samples. For example, in research and clinical applications, high-throughput genetic testing using target-specific reagents can provide accurate and precise nucleic acid target quantification for drug discovery, biomarker discovery, and clinical diagnosis. Early high-throughput determinations were performed using samples placed in microplates. However, significantly higher throughput can be obtained using emulsions and other micro-partitions. Specifically, emulsion technology can produce a large number of aqueous droplets from very small samples, which can serve as independent biochemical reaction chambers. For example, an aqueous sample (e.g., 20 microliters) can be partitioned into droplets (e.g., 20,000 droplets of 1 nanoliter each) to perform individual tests on or in each droplet.

[0003] Aqueous droplets can be suspended in oil to form water-in-oil emulsions, which are conducive to processing and analysis. Surfactants can be used to stabilize emulsions to reduce the aggregation of droplets during heating, cooling and transportation, so that thermal cycling can be performed. In some embodiments, the droplets of the emulsion are processed in a macrofluidic environment and then processed in a microfluidic environment. For example, when the droplets are in the main phase form of the emulsion, the droplets are thermally cycled in a macrofluidic environment (e.g., a sealed hole). The droplets of the emulsion are then transferred from the main phase form to a microfluidic environment (such as a droplet reader) to detect the signal from the individual droplets passing through the microfluidic channel detection zone one by one. Additional oil (exceeding the oil in the original emulsion) can be used to achieve the movement and proper spacing of droplets in a microfluidic environment. In fact, this additional oil may represent most of the oil required for analyzing the sample, or even the vast majority of the oil. Unfortunately, suitable oil may be expensive and / or difficult to obtain. Therefore, a droplet reader that can reduce the amount of oil required for analysis is needed. Summary of the invention

[0004] The present disclosure describes systems, including methods and apparatus, that use recirculated fluids to analyze partitioned samples, such as droplets. The system can be used to analyze multiple samples, wherein the fluid used for an initial sample can be reused for subsequent samples. Such reuse or recycling can reduce the amount of fluid required to perform multiple analyses while reducing costs. In addition, in some embodiments, it can simplify operations by increasing the number of analyses that can be performed before the fluid must be replenished or replaced. BRIEF DESCRIPTION OF THE DRAWINGS

[0005] Figure 1 is a high-level, generalized schematic diagram of an exemplary droplet reader according to aspects of the present disclosure.

[0006] Figure 2 is a droplet reader (such as Figure 1 Schematic diagram of existing spacer fluid and waste flow operations of a droplet reader).

[0007] Figure 3 is a schematic diagram of a first exemplary off-instrument system for recirculating spacer fluid, showing the steps and setup generated during sample analysis and subsequent oil recovery using a separatory funnel.

[0008] Figure 4 is a schematic diagram of a second exemplary off-instrument system for recirculating spacer fluid, showing the steps and setup generated during sample analysis and subsequent oil recovery using an oil separator.

[0009] Figure 5 is a schematic diagram of an exemplary on-instrument dual reservoir system for recycling spacer fluid, showing the steps and settings generated when exchanging discrete spacer fluid and waste reservoirs.

[0010] Figure 6 is a schematic diagram of a first exemplary in-instrument single reservoir system for recycling spacer fluid, showing the steps and setup generated when spacer fluid and waste share a reservoir with a single opening.

[0011] Figure 7 is a schematic diagram of a second exemplary in-instrument single reservoir system for recycling spacer fluid, showing the steps and setup generated when spacer fluid and waste share a reservoir with two openings.

[0012] Figure 8 is a side view of a first exemplary dual-port reservoir suitable for use with an in-instrument system for recirculating a spacer fluid, such as Figure 7 as shown in .

[0013] Fig. 9 is an isometric view of a second exemplary dual-port reservoir suitable for use with an in-instrument system for recirculating a spacer fluid, such as Figure 7 as shown in .

[0014] Fig.10is a comparison diagram showing representative similarities and differences when an exemplary system for recycling a spacer fluid is used with a spacer fluid that is denser than the aqueous component of the waste (upper diagram) or less dense than the aqueous component of the waste (lower diagram). In this case, the system is an embodiment of an in-instrument system in which the spacer fluid and the waste share a reservoir having two openings, such as Figure 7 shown.

[0015] Fig.11 is a schematic diagram of an exemplary inverted single reservoir system for recycling spacer fluid, which includes a catcher between the reservoir and the droplet reader.

[0016] Fig.12 is a graph showing the inverse relationship between the volume of spacer fluid in a new fluid reservoir and the number of times the spacer fluid is reused. DETAILED DESCRIPTION

[0017] The present disclosure describes systems, including methods and devices, for analyzing partitioned samples (such as droplets) using recycled fluids. The system may include a droplet reader. The droplet reader may further include (i) a sample inlet configured to receive a sample of a partition comprising an aqueous partition disposed in a carrier fluid, (ii) a spacing fluid inlet configured to input a spacing fluid for moving and / or separating the partitions, (iii) a mixing region for combining the sample and spacing fluid of the partitions, (iv) a detection region located downstream of the mixing region for interrogating the partitions, and (v) a waste outlet configured to output the sample and spacing fluid after interrogating the partitions. The droplet reader may further include at least one reservoir for storing the spacing fluid and / or receiving waste. The system may be used to analyze multiple samples, wherein the spacing fluid used for the initial sample may be reused for subsequent samples. Such reuse or recycling may reduce the amount of fluid required to perform multiple analyses while reducing costs. In addition, in some embodiments, it may simplify operation by increasing the number of analyses that can be performed before the fluid must be replenished or replaced.

[0018] Figure 1is a high-level, generalized schematic diagram of an exemplary droplet reader 20. The droplet reader includes (i) a sample inlet 22 configured to receive a partitioned sample 24 comprising an aqueous partition 26 disposed in a carrier fluid 28, (ii) a spacing fluid inlet 30 configured to input a spacing fluid 32, e.g., to move the partitions through the reader and / or increase separation between the partitions, (iii) a mixing region 34 for combining the partitioned sample and spacing fluid (such as a singulator), (iv) a detection region 36, located downstream of the mixing region, for interrogating the separated partitions, and (v) a waste outlet 38 configured to output the sample and spacing fluid (collectively referred to as "waste" or "waste fluid" 40) after the partitions have been interrogated. Typically, the partitions are aqueous fluids (such as aqueous droplets), while the carrier fluid and spacing fluid are non-aqueous fluids (such as oil) that are immiscible with the partitions. The carrier fluid and spacing fluid can be the same or different from each other. They are usually fully miscible with each other, and if they are different, they may be mostly different or completely different in the presence, absence or concentration of additives (such as surfactants). Any suitable mechanism (such as droplet generator) can be used to prepare the sample of the partition for determination, and any technology (such as polymerase chain reaction (PCR)) used to be suitable for sample and determination is processed. The sample can be an aliquot of a hole in a multi-well plate (such as a PCR flat plate or a microplate). In this case, a continuous sample can be an aliquot from the same hole or a more typical continuous hole. The spacer fluid for determination can be supplied from a reservoir (such as a discrete spacer fluid reservoir 42 accessed by a spacer fluid inlet 30). Finally, the waste 44 produced by determination can be exported to a reservoir (such as a discrete waste reservoir 44) by a waste outlet. In some embodiments, the spacer fluid and waste can be input and output from the same reservoir. In other words, the spacer fluid reservoir 42 and the waste reservoir 44 can coexist partially or completely, and the spacer fluid is extracted from a part of the reservoir, and the waste is stored and / or limited to different parts of the reservoir. The droplet reader can be placed alone, with an in-instrument or out-instrument reservoir, or with a droplet generator and / or any reaction facilitator (such as a PCR thermal cycler). The housing can protect and organize the components of the system, including the spacer fluid reservoir and the waste reservoir in this embodiment. Exemplary droplet readers, partitions, and other aspects of carriers and spacer fluids are provided in U.S. Patent Application Publication No. US-2019-0002956-A1 (now U.S. Patent No. 11,499,183), published on January 3, 2019, which is incorporated herein by reference.

[0019] Figure 2 A droplet reader (such as Figure 160 of the droplet reader of the present invention. Briefly, the spacer fluid 62 for the assay is supplied to the droplet reader (not shown) from a discrete spacer fluid reservoir 64, which is accessed via a spacer fluid inlet 66, terminated by a sedimentation filter 68, and waste 70 from the assay is output to a discrete waste reservoir 72 via a waste outlet 74 (Setup A). ​​Step 1: The assay is performed using a droplet reader (not shown), spacer fluid is drawn from the spacer fluid reservoir, and waste is output to the waste reservoir (shown as partially depleted) (Setup B). Step 2: Eventually, the spacer fluid is exhausted, and / or the waste reservoir is filled (Setup C). When the oil reservoir is low or empty and / or when the waste reservoir is partially or completely full, the reservoirs are replaced individually or in series. Importantly, the contents of the waste reservoir are discarded. In other words, in the present embodiment, the spacer fluid in the waste reservoir is not reused. Optional spacer fluid sensor 76 and waste sensor 78 may provide warnings when spacer fluid is low and / or waste is high in the respective reservoirs.

[0020] Figure 3-9 Various embodiments are shown in which a spacer fluid is recycled between sets of assays performed by a droplet reader. More specifically, a spacer fluid used in a first set of assays is reused by the droplet reader in a second set (or second set, third set, ...) of assays. To simplify the description, these embodiments may be classified (but are not limited to) as off-instrument and on-instrument embodiments. (In other words, "off-instrument embodiments" may be used on-instrument, and vice versa, as appropriate or desired. Importantly, these embodiments may be used alone or in conjunction with one another to facilitate reuse of spacer fluid from one set of assays to another. In addition, as described herein, the recycled spacer fluid may also include a carrier fluid associated with the sample that is miscible with the spacer fluid (not an aqueous phase), thereby further improving the efficacy of the embodiments. A. Out-of-instrument implementation

[0021] This section describes an exemplary extra-instrumental embodiment of a spacer fluid in the waste material output by the droplet reader for recovery and reuse. This waste material will generally be collected in a reservoir (such as a waste reservoir) to isolate and contain it until it can be processed for recovery. Suitable reservoirs include any container capable of containing (and preferably not reacting with) a liquid containing aqueous and non-aqueous components. The reservoir can have any suitable size and shape. Examples include bottles and flasks, etc., especially those that can be appropriately capped (and possibly ventilated) to reduce the possibility of contamination or leakage. The waste reservoir can be separate and different, or it can be a combined reservoir of spacer fluid and waste material (as described in the "Instrument In-Embodiment" below).

[0022] Figure 3A first exemplary off-instrument system 80 for recycling a spacer fluid is shown. Initially, the spacer fluid reservoir 82 contains a spacer fluid 84, and the waste reservoir 86 is empty (Setup A). ​​More specifically, the spacer fluid reservoir may initially be full or partially full of spacer fluid. The waste in the waste reservoir may initially be empty or partially empty. Step 1: Using a droplet reader (not shown), a determination is made that the spacer fluid is depleted, and / or the waste reservoir is filled with waste 88 (Setup B). More specifically, after use, the spacer fluid compartment may be partially empty or completely empty, and the waste reservoir may be partially or completely full. The levels of spacer fluid and / or waste in this embodiment and other embodiments can be determined using any suitable mechanism, including sensors (such as conductive sensors adjacent to the reservoir) and / or visual inspection (i.e., via the eye). Exemplary sensors may include one or more dedicated spacer fluid sensors 88 and / or one or more dedicated waste sensors 90. Step 2: Remove the waste reservoir from the instrument and pour the waste into a separatory funnel 92 (Setup C). In other embodiments, waste can be removed from the waste reservoir and added to a separatory funnel (and / or other separator) using, for example, a drain and / or siphon without removing the waste reservoir from the instrument. Step 3: Wait for an appropriate time, such as at least about 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 45 minutes, or 1 hour, etc., to allow the aqueous and non-aqueous components of the waste to separate into different spacer fluids (or spacer fluids + carrier fluids) and aqueous phases (Setting D). Step 4: The spacer fluid fraction 94a will settle to the bottom of the separatory funnel and be separated from the waste fraction 94b by opening the stopcock 95 on the separatory funnel to allow the spacer fluid to flow into the spacer fluid reservoir (or intermediate container) 82' and stopping the flow before adding the aqueous phase to the reservoir (Setting E). Step 5: The recovered spacer fluid is then returned to the same or different reservoir in the instrument for reuse (Setting A (Repeat)). Step 6: Discard the separatory funnel. Alternatively, the separatory funnel can be rinsed for reuse, for example, using bleach and ethanol, and the contents collected in a suitable reservoir 96 (Setup F). Step 7: Discard the remaining contents of the separatory funnel. Alternatively, the remaining contents can be subjected to additional separation operations, either alone or in combination with other recovered remaining contents, and then the spacer fluid is removed for use in the sample reader (Setup G).

[0023] Figure 4 A second exemplary off-instrument system 100 for recirculating a spacer fluid is shown. Figure 3 The main difference of the embodiment is that it uses an oil separator 112 (e.g., an oil separator cup) instead of a separatory funnel to separate the aqueous phase and the non-aqueous phase. Set A, B, C, D, E, F and G and steps 1, 2, 3, 4, 5, 6 and 7 are the same as Figure 3The corresponding parts in are basically the same. Here, the denser spacer fluid phase accumulates at the bottom of the cup. Then, as shown in the figure, this phase can be poured from the spout 115 on the cup into the spacer fluid reservoir 102 'or other container for reuse.

[0024] Figure 3 and 4 The separation step (step 4) shown in more generally can be performed using any suitable separation mechanism (or combination of mechanisms). Other examples can include other mechanical separators, absorption and / or distillation, etc. These mechanisms can be manual, semi-manual, semi-automatic or automatic, as appropriate or desired. The step of pouring the waste into the separation mechanism or otherwise adding the waste can include pre-filtering or otherwise pre-treating the waste, such as by absorption or skimming or otherwise extracting excess aqueous phase. The separation can be run until only a small amount of spacer fluid (such as molten metal) remains in the separation mechanism. Figure 3 and 4 As shown). Alternatively, the separation can be run until all of the spacer oil and a small amount of aqueous phase are extracted. The "spacer fluid" can then be used as is, or treated to reduce or eliminate the aqueous phase, for example by blotting or using a desiccant such as anhydrous magnesium sulfate, etc.

[0025] The separation step may be performed at any suitable location. Typically, the separation will be performed near the droplet reader so that the recovered spacing fluid can be returned to the reader for reuse. However, the separation may also be performed remotely. For example, the waste material may be shipped to another location or a third party for processing, with the aqueous phase discarded and only the recovered spacing oil returned for reuse by the same or a different user.

[0026] The method shown in the system can be used for the contents of a single waste (or combined spacer fluid and waste) reservoir. Alternatively, the contents of two, three, four, five or more reservoirs can be combined and the combined non-aqueous phases separated together. B. In-instrument implementation

[0027] This section describes exemplary in-instrument implementations for recovering and reusing spacer oil in waste output from a droplet reader. These implementations can be used alone or in combination with out-of-instrument implementations (such as those described above) to further extend the reuse of spacer fluid.

[0028] Figure 5An exemplary in-instrument system 120 for recycling oil is shown, which involves exchanging or replacing discrete spacer fluids and waste reservoirs. The exchange of reservoirs can be carried out by any suitable mechanism. For example, a depleted oil reservoir can be exchanged with a full waste reservoir, without substantially repositioning the spacer fluid input and waste output. Alternatively, the oil input and waste input can be exchanged, without substantially repositioning the spacer fluid reservoir and the waste reservoir. Initially, a full (or sufficiently full) spacer fluid reservoir 122 occupies an input position 124, connected to a spacer fluid inlet 126, and an empty (or sufficiently empty) waste reservoir 128 occupies a waste position 130, connected to a waste outlet 132 (Set A). Step 1: Using a droplet reader (not shown) to perform an assay, the volume of the spacer fluid 134 in the spacer fluid reservoir decreases as the spacer fluid is used in the assay, and the volume of the waste 135 in the waste reservoir increases as the used aqueous and non-aqueous waste is output (Set B). Step 2: The spacer fluid reservoir and waste reservoir are swapped or interchanged (Setup C). Step 3: The assay is run again, using the reservoir in the new position (or the spacer fluid inlet and waste outlet in the new position), drawing spacer fluid from the front waste reservoir, and outputting waste to the front spacer fluid reservoir. This process can be repeated until the spacer fluid reservoir is sufficiently depleted (spacer fluid) and / or the waste reservoir is sufficiently full (waste) (Setup D). Step 4: The positions of the reservoirs are switched again (Setup E). After switching, the reservoir in the spacer fluid position (or connected only to the spacer fluid inlet) is the reservoir in that position in Settings A and B, and the reservoir in the waste position is the reservoir in that position in Settings A and B (or connected only to the waste outlet). Step 5: The process can be repeated (i.e., running the assay, swapping reservoirs) until both reservoirs are full, such that no reservoir can be positioned to receive waste from the reader (Setup F). Step 6: The contents of the bottle can then be discarded, or some or all of the contents can be subjected to the procedures described in the previous section and repeated in the reader. Figure 3 and 4 One or more separate spacer fluid sensors 136 and waste sensors 138 may track the amount of spacer fluid and waste in their respective reservoirs, or simply issue a warning when spacer fluid is low and / or waste is high.

[0029] Figure 6Another exemplary in-instrument system 140 for recycling spacer fluid is shown. A single reservoir 142 with a single opening 143 is used for spacer fluid and waste material. This is possible because the aqueous components of spacer fluid and waste material are immiscible. Therefore, even if the other parts of the reservoir are occupied by the aqueous components of the waste material, the spacer fluid can be extracted from the reservoir portion occupied only by the spacer fluid. Usually, since the spacer fluid is denser than other components of water and aqueous waste material, the spacer fluid can be extracted from a lower position (below the aqueous phase) in the reservoir. In addition, since the continuous determination increases the amount of non-aqueous phase in the reservoir (because the sample contains a carrier fluid), further determination will reduce rather than increase the possibility of contamination. Initially, the reservoir 142 contains only spacer fluid 144 (setting A). The initial volume or level of the spacer fluid can be determined by a supplier or user, and is generally known. In a single reservoir embodiment, the reservoir may initially be only partially filled (such as approximately half full) to allow the accumulation of waste material combined with spacer fluid and sample and carrier fluid. The reservoir is operatively connected to a spacer fluid inlet 146 and a waste outlet 148. Step 1: Run an assay using a droplet reader (not shown), with spacer fluid 144 input from reservoir 142 through spacer fluid inlet 146 and waste 150 returned to the same reservoir through waste outlet 148. The reservoir is partially filled (Setup B). Step 2: Run additional assays, eventually filling the reservoir (Setup C). Any suitable mechanism (such as a spacer fluid sensor 152 and a waste sensor 154) can be used to determine the level of spacer fluid and waste throughout the process. The number of samples run can be determined by the initial volume of the spacer fluid and the position of the waste sensor. Alternatively or in addition, the number of samples run or the number of samples allowed to run without changing the reservoir can be tracked by software. For example, the software can require or force the user to change the reservoir after a predetermined number of runs (such as about 50 times), the specific number of times being optionally determined by the capacity of the bottle, the initial fill state of the bottle, the nature of the sample or run, the droplet reader, etc. Step 3: The spent reservoir can be discarded, or the contents of the reservoir can be processed off-instrument, as described above. Here, as described above, the reservoir has a single opening. Both the spacer fluid inlet and the waste outlet access the reservoir through the same opening. The advantage is simplicity. However, in existing dual reservoir systems (such as Figure 2 Use of a single opening reservoir in a system ) may require rearrangement of the spacing fluid inlet, waste outlet, or both.

[0030] Figure 7 Another exemplary in-instrument system 160 for recirculating spacer fluid is shown. Here, two openings 163 a , bA single reservoir 142 is used for both spacer fluid and waste. Initially, reservoir 162 contains only spacer fluid 164 (Setup A). ​​The reservoir is operatively connected to a spacer fluid inlet 166 and a waste outlet 168, wherein the inlet and outlet are each connected to a different opening. Step 1: Run an assay using a droplet reader (not shown), draw spacer fluid from the reservoir through the spacer fluid inlet, and return waste 170 to the reservoir through the waste outlet. The total level of fluid (a combination of fluid from the sample and spacer fluid from the reservoir) increases (Setup B). Step 2: When the reservoir is full or its use is complete, the contents of the reservoir can be processed as described above (with Figure 6 In particular, they can be discarded or recycled. A reservoir can include a single uninterrupted internal volume (e.g. Figure 6 ). Alternatively, as shown, the reservoir may include one or more internal partitions 172 that confine waste to only a portion of the surface area of ​​the fluid in the reservoir. This may make it easier to extract uncontaminated spacer fluid and / or reduce contamination of the spacer fluid by reducing the interface between the spacer fluid and the waste. Here, as described above, the reservoir has separate openings for receiving a spacer fluid input and a waste outlet. These openings may have any suitable size and shape. In the illustrated embodiment, the openings have the same size as the openings on the discrete spacer fluid and waste reservoirs, and have the same position and distance within the openings as the specific reservoir when installed for use in the droplet reader. Thus, the droplet reader may alternatively and conveniently be used with discrete spacer fluid and waste reservoirs and combined spacer fluid and waste reservoirs.

[0031] Figure 8 A first exemplary dual-port partitioned reservoir 180 suitable for use with an in-instrument system for recirculating a spacer fluid, such as Figure 7 The reservoir 180 has two openings 181 a , b These openings may receive spacer fluid inlets and waste outlets, such as Figure 7 The storage also includes partitions 182, such as Figure 7 The reservoir also includes an opening 181 a , b Asymmetric bottom 184 below a , b Therefore, in the illustrated embodiment, the opening 181 a The depth of the reservoir directly below (or accessible fluid depth) (denoted as D a ) is larger than opening 181 b The depth of the reservoir directly below (or accessible fluid depth) (denoted as D b ). Therefore, through the opening 181a The positioned spacer fluid inlet can be positioned deeper in the reservoir and then through the remaining opening 181 b The waste material of the reservoir is further below, making it more likely that the spacer fluid can be input without being contaminated by the waste material. The reservoir can also include a gap 184, which can exclude fluid while giving the reservoir a flat bottom, thereby facilitating stable placement in the droplet reader. The gaps and partitions can further define and reduce the total volume of the reservoir. The reduced volume can reduce the total number of measurements that can be performed on the reservoir before the reservoir is full and must be replaced. This in turn can reduce the time between reservoir changes, making the contents of the reservoir less likely to be contaminated, for example, due to fungal and / or bacterial growth in the reservoir. Finally, the spacing or width (denoted as W) from opening to opening can be selected to match the spacing from opening to opening of a single discrete reservoir in the droplet reader, thereby promoting interchangeability.

[0032] Fig. 9 A second exemplary dual-port partitioned reservoir 190 suitable for use with an in-instrument system for recirculating a spacer fluid, such as Figure 7 As shown. The storage 190 and Figure 8 The reservoir 180 in FIG. 1 has many similarities and can be used in the same manner and to obtain the same benefits. For example, like the reservoir 180, the reservoir 190 has two openings 191 a , b and an intermediate partition 192 for limiting output waste. It may also have an opening-to-opening spacing or width (denoted as W') that matches the opening-to-opening spacing of a single discrete reservoir used in the same droplet reader. Reservoir 190 may have a constant depth or an asymmetric depth. Reservoir 190 may be symmetrical, or at least substantially symmetrical, across two openings 191. a , b The center plane P1 or perpendicular to the plane P1 and the opening 191 a , b equidistant plane P2. Alternatively, reservoir 190 may be asymmetrical about one or both of these planes. For example, a reservoir may include a cutout 195 on one side but not on the other side. The cutout may be used for any suitable purpose, such as an alignment marker (e.g., to ensure that the reservoir is properly positioned in a droplet reader), a volume adjuster (e.g., to reduce the volume of one side of the reservoir relative to the other side), and / or a void for receiving a label, sensor, and / or tag (e.g., an RFID tag), etc. The cutout may similarly have any suitable size and location consistent with its intended function.

[0033] As used herein, a reservoir may have any suitable size, shape, and number of openings. A reservoir may include a single, unpartitioned volume. Alternatively, Figure 7-9As shown, the reservoir may include a partition or separator to divide the upper portion of the volume. A denser spacer fluid may flow between the two (or more) portions by traveling under the separator. However, the aqueous portion of the waste material floating on the spacer fluid is confined to only that portion of the volume.

[0034] The carrier and spacing fluids used herein can have any suitable composition, density, and / or other physical properties consistent with their use in creating, separating, and moving partitioned samples. Examples include fluorinated oils, silicone oils, and hydrocarbon oils, among others. In some cases, such as Figure 2-9 In those shown in , the spacer fluid is denser than water, so that when the two come into contact, the aqueous component of the waste floats on the spacer fluid. Exemplary denser spacer fluids can include fluorinated oils, etc. In other cases, the spacer fluid may be less dense than water, so that the spacer fluid floats on the aqueous component rather than sinking below the aqueous component. Exemplary less dense spacer fluids can include silicone oil and mineral oil, etc. The systems described herein will work with denser or less dense spacer fluids. For example, Figure 3 and 4 The off-instrument separation mechanism in can be used "in reverse" to collect the upper rather than the lower fraction. Figure 3 In the case of using a separatory funnel, this can be achieved by first discharging the lower aqueous fraction, optionally including a small amount of the upper fraction, and then collecting the upper spacer fluid fraction for reuse. Figure 4 In the embodiment of the present invention, using an oil separator, the lower aqueous portion can be decanted, optionally including a small amount of the upper fraction, and then the remaining upper fraction can be collected for reuse. A similar method can be used for in-instrument embodiments, taking care to draw the spacer fluid from the upper rather than the lower fraction. This method is particularly suitable for single-reservoir embodiments such as Figure 6 and 7 Those in which the spacer fluid inlet is located high rather than low within the reservoir. This approach is particularly useful in dual-opening single-reservoir embodiments where partitions in the fluid volume can help separate the spacer fluid and aqueous components of the waste. Fig.10 is a comparison of such a system 200 in which the spacer fluid is denser (upper figure) or less dense (lower figure) than the aqueous components of the waste. The waste components floating on the spacer fluid in the upper figure sink below the spacer fluid in the lower figure. Thus, in order to confine the waste 202 to one side of the reservoir 204 below the waste outlet, the portion of the partition 206 that contacts the waste can be from the top of the upper figure downward and from the bottom of the lower figure upward. The gaps or holes 210 in the partition can be relatively low in the upper figure and relatively high in the lower figure, in both cases allowing the spacer fluid 212 to move throughout the reservoir while preferentially confining the waste to one side. C. Inverted embodiment (including trap)

[0035] This section describes an exemplary inverted embodiment of the spacer oil in the waste material outputted by the drop reader for recovery and reuse. In these embodiments, the spacer fluid input and / or waste material output can access the corresponding spacer fluid and waste material reservoir or shared reservoir through the bottom of the reservoir rather than the top. Here, "bottom" refers to the point below the fluid level in the reservoir, and is generally located at the lowest point or near the reservoir. Fluid can be pumped into and / or pumped out of the reservoir. In some cases, the fluid may leave the reservoir (i.e., by gravity feeding) due to gravity. The reservoir shown here and / or in the previous section can combine the trap, for example, as described below.

[0036] Fig.11 An exemplary inverted system 220 for recycling spacer fluid is shown. Here, a single inverted reservoir 222 is used to hold spacer fluid 224 for use by a droplet reader 226 and to receive waste 228 generated by the droplet reader. The reservoir and droplet reader are separated by an intermediate catcher 230 fluid, which may contain additional spacer fluid 232 and / or waste 234. The spacer fluid flows (e.g., under gravity) and / or is pumped from the catcher to the droplet reader for use in the droplet reader as needed. During and / or after droplet reading, the waste flows and / or is pumped back to the catcher from the droplet reader. The illustrated embodiment is intended to be used when the spacer fluid is denser than the non-recirculated (e.g., aqueous or sample) portion of the waste so that the waste floats on the spacer fluid. In this case, the denser spacer fluid leaves the trap through a trap spacer fluid outlet 236 located at or near the bottom 238 of the trap, while waste (including sample, carrier fluid, and spacer fluid) enters the trap through a trap waste inlet 240 located at or near the top 242 of the trap. The trap is in fluid communication with the reservoir through a common fluid channel 244 between a trap port 246 located at the top of the trap and a reservoir port 248 located at the bottom of the reservoir. The system may also include a vent 250 and / or a sensor 252. a , b , for monitoring compartment fluid, waste, and / or total fluid volume in the reservoir and / or trap. Alternatively, or in addition, to track fluid volume, the system may include a software feature to calculate volume based on the number of runs performed and the typical volume of fluid used and generated per run. The bottle may include one or more identifiers, such as a serial number, a bar code, and / or a radio frequency identification (RFID) tag, etc. In this case, the software may prevent a user from reusing a particular bottle on a particular machine after a predetermined limit (such as the number of runs) is reached.

[0037] The system can be used as follows. Step 1: Connect a reservoir with a spacer fluid to the system via a base 254 and allow the spacer fluid to flow from the reservoir into the trap through a common fluid channel. Step 2: Move the spacer fluid from the trap to an onboard spacer fluid reservoir 256 on the droplet reader via the trap spacer fluid outlet. Step 3: The droplet reader runs a read cycle, mixes the spacer fluid with the partitioned sample (droplets and carrier fluid) 258, detects signals from the droplets, and generates waste that is stored in a tank 260 (such as a coil in a pipe or other intermediate reservoir on the droplet reader). If desired, this step can be repeated with multiple samples until the onboard reservoir is exhausted. Step 4: Pump the waste contained in the tank back to the trap via the trap waste inlet. Alternatively, or in addition, in some embodiments, waste can be continuously moved to the trap during operation of the droplet reader. The waste material moves upward into the reservoir through the common channel, the spacer fluid stays or settles to the bottom, and the less dense components of the waste material (specifically the sample) float to the top. If necessary, the common fluid line can be flushed to remove residual waste material. Step 5: The above steps (specifically steps 2-4) can be repeated until the reservoir is full (due to the addition of waste material and carrier fluid). The reservoir can then be replaced (step 1) and the process repeated.

[0038] The reservoir and / or trap may be located within the instrument or outside the instrument. The reservoir may be used in a system configured for a single reservoir or in a system configured for two (or more) reservoirs but modified for use with a single reservoir. In the latter case, the trap may be added and the additional reservoir base 262 may be taken offline (e.g., by inserting a bypass valve 264 and closing the line from the additional base to the reservoir using a shutoff valve 266 or other suitable mechanism). D. Quantification of compartment fluid recirculation

[0039] This section explores the quantitative aspects of interval fluid recirculation.

[0040] Fig.12 is a graph showing the inverse relationship between the volume of isolation fluid in a fresh fluid reservoir, the number of samples analyzed per reservoir (measured in "plates"), and the number of times the isolation fluid is reused. E. Illustrative combinations and additional examples

[0041] This section describes additional aspects and features of the systems, methods, and devices of the present disclosure, presented as, but not limited to, a series of paragraphs, some or all of which may be alphanumerically indexed for clarity and efficiency. Each of these paragraphs may be combined in any suitable manner with one or more other paragraphs, and / or with the disclosure elsewhere in the application. Some of the paragraphs below explicitly mention and further limit other paragraphs, providing examples of some suitable combinations, but are not limited thereto.

[0042] A. A method for analyzing multiple samples, comprising: (1) providing a sample reader having (i) a sample inlet configured to receive a sample of a partition comprising an aqueous partition disposed in a carrier fluid, (ii) a spacer fluid inlet configured to input a spacer fluid, (iii) a mixing region for combining the sample of the partition and the spacer fluid, (iv) a detection region located downstream of the mixing region for interrogating the partition, and (v) a waste outlet configured to output the sample and the spacer fluid after the partition is interrogated; (2) loading a first sample of the partition into the sample reader, and (1) inputting a spacer fluid through the sample inlet, combining the sample and the spacer fluid in the mixing zone, interrogating the partitions in the detection zone, and collecting waste containing the first sample and the associated spacer fluid from the waste outlet after the first sample is interrogated; and (2) loading a partitioned second sample into the sample reader, inputting a spacer fluid through the spacer fluid inlet, combining the sample and the spacer fluid in the mixing zone, interrogating the partitions in the detection zone, and collecting waste containing the second sample and the associated spacer fluid from the waste outlet after the second sample is interrogated; wherein at least a portion of the spacer fluid combined with the second sample has been recovered from the spacer fluid combined with the first sample.

[0043] A1. The method of paragraph A, wherein the partitions are droplets.

[0044] A2. The method of paragraph A or A1, wherein the spacer fluid is oil.

[0045] A3. The method according to paragraph A2, wherein the oil is selected from the following group: fluorinated oil, silicone oil and hydrocarbon oil.

[0046] A4. A method according to any of the preceding paragraphs, wherein the carrier fluid and the spacer fluid are miscible.

[0047] A5. The method of paragraph A4, wherein the carrier fluid and the spacer fluid differ only in their respective additives.

[0048] A6. The method according to any of the preceding paragraphs, further comprising dividing the first sample into a plurality of partitions separated by a carrier fluid to form a partitioned first sample, and dividing the second sample into a plurality of partitions separated by a carrier fluid to form a partitioned second sample.

[0049] A7. The method according to any one of the preceding paragraphs, further comprising selecting a droplet generator, wherein the step of dividing the first sample and the step of dividing the second sample are performed using a droplet generator, wherein the multiple partitions of the first sample and the multiple partitions of the second sample are droplets.

[0050] A8. According to the method described in paragraph A6 or A7, the first sample and the second sample contain nucleic acid, which further includes amplifying the nucleic acid after the step of dividing the first and second samples into a plurality of partitions and before the step of loading the partitioned samples into a sample reader.

[0051] A9. A method according to any of the preceding paragraphs, wherein the step of interrogating the partitions involves determining the number of partitions that are positive for nucleic acid amplification.

[0052] A10. The method of any of the preceding paragraphs, wherein the step of interrogating the partition comprises measuring fluorescence emission from the partition.

[0053] A11. A method according to any of the preceding paragraphs, wherein the mixing region is a disperser configured to increase the separation between upstream partitions of the detection region.

[0054] A12. According to the method described in any of the preceding paragraphs, the sample reader further has at least one reservoir configured to contain a spacing fluid and waste, wherein the spacing fluid inlet is arranged to input the spacing from the at least one reservoir, and the waste outlet is arranged to output the waste to the at least one reservoir.

[0055] A13. According to the method described in paragraph A12, the sample reader further includes at least one of a spacing fluid sensor and a waste sensor, and the spacing fluid sensor and the waste sensor are configured to determine the level of spacing fluid or waste in the at least one reservoir, respectively. One-Reservoir Implementation

[0056] A14. According to the method described in paragraph A12 or A13, at least one reservoir includes a combined reservoir for a spacer fluid and waste, wherein the spacer fluid and the sample are immiscible, and wherein the spacer fluid inlet extracts the spacer fluid from the portion of the combined reservoir occupied by the spacer fluid.

[0057] A15. The method of paragraph A14, wherein the sample has a density less than the spacer fluid such that the sample floats on the spacer fluid, and wherein the input of the spacer fluid is arranged in the spacer fluid below the level of the sample in the combined reservoir.

[0058] A15A. The method of paragraph A14, wherein the sample has a density greater than the spacer fluid such that the spacer fluid floats on the sample, and wherein the input of the spacer fluid is arranged in the spacer fluid above the level of the sample in the combination reservoir.

[0059] A16. The method of any of paragraphs A14 to A15A, wherein the combined reservoir has a single opening, and wherein the spacer fluid inlet and the waste outlet both access the combined reservoir through the same opening.

[0060] A17. A method according to any of paragraphs A14 to A15A, wherein the combined reservoir has two openings, and wherein the spacer fluid inlet accesses the combined reservoir through one opening, and wherein the waste outlet accesses the combined reservoir through the other opening.

[0061] A18. A system according to paragraph A17, wherein the sample reader is configured for use with a combined spacer fluid and waste reservoir or with a discrete spacer fluid reservoir and a discrete waste reservoir, and wherein when the discrete reservoirs are properly positioned for use, the separation between the openings in the combined reservoir is the same as the separation between the openings in the discrete reservoirs.

[0062] A19. The method of paragraph A17 or A18, wherein each opening is connected to a common volume by a neck, and one of the necks is wider than the other neck.

[0063] A20. According to the method described in any one of paragraphs A14 to A19, the sample reader further has at least one sensor connected to the combination reservoir, wherein the sensor is configured to report at least one of the spacer fluid level, the waste level, and the total level of the spacer fluid and waste in the combination reservoir.

[0064] A21. A method according to any of paragraphs A14 to A20, wherein the volume of the combined reservoir is selected to limit the amount of time it takes for the bottle to be filled to reduce the likelihood that the waste material will deteriorate before the bottle is filled.

[0065] A22. The method of any of paragraphs A14 to A21, wherein the volume of the combined reservoir is less than or equal to about 1500 mL.

[0066] A23. The method of paragraph A22, wherein the volume of the combined reservoir is about 1000 mL. Two-Reservoir Implementation

[0067] A24. According to the method described in paragraph A12 or A13, the at least one reservoir includes a discrete interval fluid reservoir and a discrete waste reservoir, wherein the interval fluid inlet is arranged to input intervals from the interval fluid reservoir, and the waste outlet is arranged to output waste into the waste reservoir.

[0068] A25. The method according to paragraph A24 further includes exchanging the spacer fluid reservoir and the waste reservoir between the steps of loading the first sample and loading the second sample, so that the spacer fluid to be combined with the second sample comes from the waste reservoir, and the waste reservoir is used to receive the sample and spacer fluid from the first sample.

[0069] A26. The method according to paragraph A25 further includes: (1) loading a partitioned third sample into a sample reader, inputting a spacer fluid through a spacer fluid inlet, combining the sample and the spacer fluid in a mixing area, interrogating the partition in a detection area, and collecting the combined third sample and spacer fluid from a waste outlet after the third sample is interrogated; and (2) exchanging the spacer fluid reservoir and the waste reservoir between the steps of loading the second sample and loading the third sample, so that the spacer fluid combined with the third sample comes from the waste reservoir, which is used to receive the sample and spacer fluid from the second sample.

[0070] A27. The method of paragraph A25 or A26, further comprising exchanging the spacer fluid reservoir and the waste reservoir until at least one reservoir is filled with sample and spacer fluid and cannot be reused without reducing its contents. Out-of-instrument implementation

[0071] A28. The method of any of paragraphs A12 to A27, wherein the waste comprises a spacer fluid and an aqueous component, further comprising separating the spacer fluid from the aqueous component and reusing the spacer fluid to analyze additional samples.

[0072] A29. The method of paragraph A28, wherein the step of separating the spacer fluid from the aqueous component is performed using at least one of a separatory funnel and an oil separator.

[0073] A30. The method according to paragraph A29 further includes transferring the contents of the waste reservoir to a separatory funnel, waiting for the contents to separate into used sample and used spacer fluid, collecting the used spacer fluid from the separatory funnel, and adding the used spacer fluid to the spacer fluid reservoir for reuse in the sample reader.

[0074] A31. The method according to paragraph A29 further includes transferring the contents of the waste reservoir to an oil separator, waiting for the contents to separate into used sample and used spacer fluid, collecting the used spacer fluid from the oil separator, and adding the used spacer fluid to the spacer fluid reservoir for reuse in the sample reader.

[0075] A32. A method according to any of paragraphs A28-A31, further comprising transporting waste to a remote location, separating the spacing fluid from the aqueous component at the remote location, and transporting the separated spacing fluid back for use with the same or a different droplet reader. Inverted and / or Trap Embodiments

[0076] A33. According to the method described in any one of paragraphs A-A11, the sample reader further has a reservoir and a trap that are fluidically connected to each other, and the reservoir and the trap are each configured to contain a spacing fluid and waste, and further includes extracting the spacing fluid used to interrogate the sample from the trap and returning the waste generated by the interrogation sample to the trap.

[0077] A34. The method of paragraph A33, further comprising transferring waste from the collector to the reservoir.

[0078] A35. The method of paragraph A33 or A34, further comprising drawing spacer fluid from the reservoir into the trap to replenish the supply of spacer fluid in the trap.

[0079] A36. The method of any of paragraphs A33-A35, wherein waste material is transferred from the trap to the reservoir and a spacer fluid is drawn from the reservoir to the trap using a common fluid path.

[0080] A37. The method of paragraph A36, wherein the common fluid path connects the top of the trap to the bottom of the reservoir.

[0081] A38. The method of any of paragraphs A33-A37, further comprising transferring fluid from the catcher to an onboard reservoir on the droplet reader.

[0082] A39. The method of paragraph A38, wherein the transferring step involves gravity feeding.

[0083] A40. The method of paragraph A38 or A39, wherein the spacer fluid used to interrogate the first and second samples is obtained from an onboard reservoir without replenishment from a trap.

[0084] A41. The method of any of paragraphs A38-A40, further comprising transferring additional spacer fluid from the trap to the onboard reservoir after interrogating the first and second samples or after the onboard reservoir has been exhausted, and then interrogating additional samples.

[0085] A42. A method according to any of paragraphs A33-A41, wherein the first and second samples are interrogated and waste is generated without returning the waste generated by the interrogation to the trap.

[0086] A43. The method of paragraph A42, wherein the waste generated by interrogation is stored in an online holding reservoir and transferred to the trap after interrogation of the first and second samples.

[0087] A44. A method according to any of the preceding paragraphs, wherein a software feature tracks the number of samples interrogated, and wherein the software feature prevents analysis of additional samples after a preselected number of samples have been interrogated.

[0088] B. A system for analyzing multiple samples, comprising: (1) a sample reader having (i) a sample inlet configured to receive a sample comprising an aqueous partition arranged in a carrier fluid, (ii) a spacer fluid inlet configured to input a spacer fluid, (iii) a mixing region for combining the sample of the partition and the spacer fluid, (iv) a detection region located downstream of the mixing region for interrogating the partition, and (v) a waste outlet configured to output the sample and the spacer fluid after the partition is interrogated; and (2) a combined spacer fluid and waste reservoir configured to hold the spacer fluid for input into the sample reader through the spacer fluid inlet and to receive waste comprising the sample and the associated spacer fluid from the sample reader through the waste outlet, wherein the spacer fluid and the waste are in contact in the reservoir.

[0089] B1. A system according to paragraph B, further comprising (a) at least one sensor configured to report at least one of a spacer fluid level, a waste level, and a total level of spacer fluid and waste in a combined reservoir, and / or (b) a software feature for tracking the number of samples that have been interrogated from the combined reservoir and optionally requiring replacement of the reservoir after a predetermined number of samples have been interrogated.

[0090] B2. A system according to paragraph B or B1, wherein the combined reservoir has a single opening, and wherein the spacer fluid inlet and the waste outlet both enter the combined reservoir through the single opening.

[0091] B3. A system according to paragraph B or B1, wherein the combined reservoir has two openings, and wherein the spacer fluid inlet accesses the combined reservoir through one opening, and wherein the waste outlet accesses the combined reservoir through the other opening.

[0092] B4. A system according to paragraph B3, wherein the combined reservoir includes at least one partition that confines waste to only a portion of the total fluid surface within the combined reservoir, and wherein the portion is at least partially located below the waste outlet.

[0093] B5. A system according to paragraph B3 or B4, further comprising a discrete spacing fluid reservoir configured to contain a spacing fluid for input into the sample reader through a spacing fluid inlet, and a discrete waste reservoir configured to receive waste from the sample reader through a waste outlet, wherein the sample reader is configured to interchangeably use a combined spacing fluid and waste reservoir or a discrete spacing fluid reservoir and a discrete waste reservoir to simultaneously analyze a sample.

[0094] B6. A system as described in paragraph B5, wherein the separation between the openings in the combination reservoir is the same as the separation between the openings in the discrete spacer fluid reservoir and the discrete waste reservoir when the discrete reservoirs are properly positioned for use with a sample reader.

[0095] B7. A system according to any of paragraphs B3 to B6, wherein each opening is connected to a common volume by a neck, and wherein the neck associated with one opening is wider than the neck associated with another opening.

[0096] B8. A system as described in any of paragraphs B3 to B7, wherein the accessible fluid depth directly beneath one opening is different from the accessible fluid depth directly beneath another opening.

[0097] B9. A system according to paragraph B8, wherein the spacer fluid is optionally denser than the sample, wherein the spacer fluid inlet accesses the reservoir through an opening associated with a larger accessible fluid depth, and the waste outlet accesses the reservoir through an opening associated with a smaller accessible fluid depth.

[0098] B10. The system of paragraph B8 or B9, wherein the combination reservoir has a bottom configured to rest on a surface when the reservoir is in use, wherein each opening is the same distance from the bottom.

[0099] B11. A system according to paragraph B or B1, wherein the combined reservoir has a single opening and wherein a trap fluid configured to hold additional spacer fluid and waste is inserted between the reservoir and the sample reader.

[0100] B12. A system according to paragraph B11, wherein the sample reader receives spacing fluid from the trap and returns waste to the trap.

[0101] B13. A system according to paragraph B12, wherein the spacer fluid and the waste material are exchanged with fluid in the reservoir so that the waste material moves from the trap to the reservoir and the spacer fluid moves from the reservoir to the trap.

[0102] B14. A system according to any of paragraphs B11-B13, wherein the sample reader further has an onboard reservoir for holding a spacing fluid received from the trap, and at least one waste tank for holding waste before it is returned to the tank.

[0103] B15. A system according to any of paragraphs B to B14, wherein the partitions are droplets, and the system further includes a droplet generator configured to divide the sample into droplets before the sample is analyzed by the sample reader.

[0104] B16. The system of paragraph B15, further comprising a thermal cycler configured to thermally cycle the droplets after the droplet generator generates the droplets and before the sample reader analyzes the droplets.

[0105] B17. A system according to any of paragraphs B to B16, wherein the volume of the combined reservoir is less than or equal to about 1500 mL.

[0106] B18. A system according to paragraph B17, wherein the volume of the combined reservoir is about 1000 mL.

[0107] B19. The system of any of paragraphs B to B18, wherein the spacer fluid has a greater density than the sample, and wherein the spacer fluid inlet is positioned toward the bottom of the combination reservoir.

[0108] B20. A system according to any of paragraphs B to B18, wherein the sample has a greater density than the spacer fluid, and wherein the spacer fluid inlet is positioned toward the top of the combined reservoir.

[0109] The above disclosure may include multiple different inventions with independent utility. Although various of these inventions have been disclosed in their preferred forms, the specific embodiments of the invention disclosed and illustrated herein should not be considered restrictive, as many changes can be made. The subject matter of the present invention includes all novel and non-obvious combinations and sub-combinations of the various elements, features, functions and / or characteristics disclosed herein. The claims specifically point out certain combinations and sub-combinations that are novel and non-obvious. Inventions embodied in other combinations and sub-combinations of features, functions, elements and / or characteristics may be claimed in applications claiming priority to the present application or a related application. Such paragraphs, whether for different inventions or the same invention, and wider, narrower, the same or different in scope than the original paragraph, are also deemed to be included in the subject matter of the invention disclosed herein.

Claims

1. A method for analyzing a plurality of samples, comprising: A sample reader is provided having (i) a sample inlet configured to receive a sample of a partition comprising an aqueous partition disposed in a carrier fluid, (ii) a spacing fluid inlet configured to input a spacing fluid, (iii) a mixing region for combining the sample of the partition and the spacing fluid, (iv) a detection region located downstream of the mixing region for interrogating the partition, and (v) a waste outlet configured to output the sample and the spacing fluid after interrogating the partition; loading a first sample of the partition into a sample reader, inputting a spacing fluid through a spacing fluid inlet, combining the sample with the spacing fluid in a mixing region, interrogating the partition in a detection region, and collecting waste comprising the first sample and associated spacing fluid from a waste outlet after interrogating the first sample; and loading a partitioned second sample into a sample reader, inputting a spacer fluid through a spacer fluid inlet, combining the sample and the spacer fluid in a mixing region, interrogating the partition in a detection region, and collecting waste comprising the second sample and associated spacer fluid from a waste outlet after interrogating the second sample; wherein at least a portion of the spacer fluid combined with the second sample has been recovered from the spacer fluid combined with the first sample. The method of claim 1 , wherein the partitions are droplets.

3. The method of claim 1, wherein the spacer fluid is oil.

4. The method according to claim 3, wherein the oil is selected from the group consisting of fluorinated oils, silicone oils and hydrocarbon oils.

5. The method of claim 1, wherein the carrier fluid and the spacer fluid are miscible.

6. The method of claim 5, wherein the carrier fluid and the spacer fluid differ only in their respective additives.

7. The method of claim 1, further comprising partitioning the first sample into a plurality of partitions separated by a carrier fluid to form a partitioned first sample, and partitioning the second sample into a plurality of partitions separated by a carrier fluid to form a partitioned second sample.

8. The method of claim 1, further comprising selecting a droplet generator, wherein: The step of partitioning the first sample and the step of partitioning the second sample are performed using a droplet generator, wherein the plurality of partitions of the first sample and the plurality of partitions of the second sample are droplets.

9. The method according to claim 7, wherein the first sample and the second sample contain nucleic acid, further comprising amplifying the nucleic acid after the step of dividing the first and second samples into a plurality of partitions and before the step of loading the partitioned samples into a sample reader.

10. The method of claim 1, wherein the step of interrogating the partitions involves determining the number of partitions that are positive for nucleic acid amplification.

11. The method of claim 1 , wherein the step of interrogating the partition comprises measuring fluorescence emission from the partition.

12. The method of claim 1, wherein the mixing region is a disperser configured to increase separation between zones upstream of the detection region.

13. The method of claim 1, wherein the sample reader further has at least one reservoir configured to contain a spacer fluid and waste, wherein a spacer fluid inlet is arranged to input a spacer from the at least one reservoir, and a waste outlet is arranged to output waste to the at least one reservoir.

14. The method of claim 13, the sample reader further comprising at least one of a spacer fluid sensor and a waste sensor configured to determine a level of spacer fluid or waste, respectively, in the at least one reservoir.

15. The method of claim 13, wherein at least one reservoir comprises a combined reservoir for spacer fluid and waste, wherein the spacer fluid and the sample are immiscible, and wherein the spacer fluid inlet extracts the spacer fluid from the portion of the combined reservoir occupied by the spacer fluid.

16. The method of claim 15, wherein the sample has a density less than the spacer fluid such that the sample floats on the spacer fluid, and wherein the input of the spacer fluid is arranged in the spacer fluid below the level of the sample in the combined reservoir.

17. The method of claim 15, wherein the sample has a density greater than the spacer fluid such that the spacer fluid floats on the sample, and wherein the input of the spacer fluid is arranged in the spacer fluid above the level of the sample in the combined reservoir.

18. The method of claim 15, wherein the combined reservoir has a single opening, and wherein the spacer fluid inlet and the waste outlet both access the combined reservoir through the same opening.

19. The method of claim 15, wherein the combination reservoir has two openings, and wherein the spacer fluid inlet opens into the combination reservoir through one opening, and wherein the waste outlet opens into the combination reservoir through the other opening.

20. A system according to claim 19, wherein the sample reader is configured for use with a combined spacer fluid and waste reservoir or with a discrete spacer fluid reservoir and a discrete waste reservoir, and wherein the separation between the openings in the combined reservoir is the same as the separation between the openings in the discrete reservoirs when the discrete reservoirs are properly positioned for use.

21. The method of claim 19, wherein each opening is connected to the common volume by a neck, and wherein one of the necks is wider than the other neck.

22. The method of claim 15, wherein the sample reader further has at least one sensor in communication with the combination reservoir, wherein the sensor is configured to report at least one of a spacer fluid level, a waste level, and a total level of spacer fluid and waste in the combination reservoir.

23. The method of claim 15, wherein the volume of the combined reservoir is selected to limit the amount of time it takes for the bottle to be filled to reduce the likelihood that the waste material will deteriorate before the bottle is filled.

24. The method of claim 15, wherein the volume of the combined reservoir is less than or equal to about 1500 mL.

25. The method of claim 24, wherein the volume of the combined reservoir is about 1000 mL.

26. The method of claim 13, wherein the at least one reservoir comprises a discrete interval fluid reservoir and a discrete waste reservoir, wherein the interval fluid inlet is arranged to input intervals from the interval fluid reservoir, and the waste outlet is arranged to output waste into the waste reservoir.

27. The method of claim 26, further comprising exchanging a spacer fluid reservoir and a waste reservoir between the steps of loading a first sample and loading a second sample, so that the spacer fluid to be combined with the second sample comes from the waste reservoir, which is used to receive the sample and the spacer fluid from the first sample.

28. The method of claim 27, further comprising: loading the partitioned third sample into a sample reader, inputting a spacer fluid through a spacer fluid inlet, combining the sample with the spacer fluid in a mixing region, interrogating the partition in a detection region, and collecting the combined third sample and spacer fluid from a waste outlet after interrogating the third sample; and The spacer fluid reservoir and the waste reservoir are exchanged between the steps of loading the second sample and loading the third sample so that the spacer fluid combined with the third sample comes from the waste reservoir that is used to receive the sample and spacer fluid from the second sample.

29. The method of claim 27, further comprising exchanging spacer fluid reservoirs and waste reservoirs until at least one reservoir is so full of sample and spacer fluid that it cannot be reused without reducing its contents.

30. The method of claim 13, the waste comprising a spacer fluid and an aqueous component, further comprising separating the spacer fluid from the aqueous component and reusing the spacer fluid to analyze additional samples.

31. The method of claim 30, wherein the step of separating the spacer fluid from the aqueous component is performed using at least one of a separatory funnel and an oil separator.

32. The method of claim 31 , further comprising transferring the contents of a waste reservoir to a separatory funnel, waiting for the contents to separate into a used sample and a used spacer fluid, collecting the used spacer fluid from the separatory funnel, and adding the used spacer fluid to a spacer fluid reservoir for reuse in a sample reader.

33. The method of claim 31 , further comprising transferring the contents of a waste reservoir to an oil separator, waiting for the contents to separate into a used sample and a used spacer fluid, collecting the used spacer fluid from the oil separator, and adding the used spacer fluid to a spacer fluid reservoir for reuse in a sample reader.

34. The method of claim 30, further comprising transporting waste to a remote location, separating the spacer fluid from the aqueous component at the remote location, and transporting the separated spacer fluid back for use with the same or a different droplet reader.

35. The method of claim 1, wherein the sample reader further comprises a reservoir and a trap in fluid communication with each other, wherein the reservoir and the trap are each configured to contain a spacer fluid and waste, and further comprising extracting a spacer fluid for interrogating a sample from the trap and returning waste generated by the interrogation of the sample to the trap.

36. The method of claim 35, further comprising transferring waste material from the trap to the reservoir.

37. The method of claim 35, further comprising drawing spacer fluid from the reservoir into the trap to replenish the supply of spacer fluid in the trap.

38. The method of claim 35, wherein waste material is transferred from the trap to the reservoir and spacer fluid is drawn from the reservoir to the trap using a common fluid path.

39. The method of claim 38, wherein the common fluid path connects the top of the trap to the bottom of the reservoir.

40. The method of claim 35, further comprising transferring fluid from the catcher to an onboard reservoir on the droplet reader.

41. The method of claim 40, wherein the transferring step involves gravity feeding.

42. The method of claim 40, wherein the spacer fluid used to interrogate the first and second samples is obtained from an onboard reservoir without replenishment from a trap.

43. The method of claim 40, further comprising transferring additional spacer fluid from the trap to the onboard reservoir after interrogating the first and second samples or after an onboard reservoir has been depleted, and then interrogating additional samples.

44. The method of claim 35, wherein the first and second samples are interrogated and waste is generated without returning waste generated by the interrogation to a trap.

45. The method of claim 44, wherein said waste material generated by interrogation is stored in an in-line holding reservoir and transferred to said trap after interrogation of said first and second samples.

46. ​​The method of claim 1, wherein a software feature tracks the number of samples interrogated, and wherein the software feature prevents analysis of additional samples after a preselected number of samples have been interrogated.

47. A system for analyzing a plurality of samples, comprising: A sample reader having (i) a sample inlet configured to receive a sample comprising an aqueous partition disposed in a carrier fluid, (ii) a spacing fluid inlet configured to input a spacing fluid, (iii) a mixing region for combining the sample of the partition and the spacing fluid, (iv) a detection region located downstream of the mixing region for interrogating the partition, and (v) a waste outlet configured to output the sample and the spacing fluid after the partition has been interrogated; A combined spacer fluid and waste reservoir is configured to contain a spacer fluid for input into a sample reader through a spacer fluid inlet and to receive waste comprising a sample and an associated spacer fluid from the sample reader through a waste outlet, wherein the spacer fluid and waste are in contact in the reservoir.

48. The system of claim 47, further comprising (a) at least one sensor configured to report at least one of a spacer fluid level, a waste level, and a total level of spacer fluid and waste in a combined reservoir, and / or (b) a software feature for tracking the number of samples that have interrogated the combined reservoir and optionally requiring replacement of the reservoir after a predetermined number of samples have been interrogated.

49. The system of claim 47, wherein the combined reservoir has a single opening, and wherein the spacer fluid inlet and the waste outlet both access the combined reservoir through the single opening.

50. The system of claim 47, wherein the combination reservoir has two openings, and wherein the spacer fluid inlet opens into the combination reservoir through one opening, and wherein the waste outlet opens into the combination reservoir through the other opening.

51. A system according to claim 50, wherein the combined reservoir includes at least one partition that confines waste to only a portion of the total fluid surface within the combined reservoir, and wherein the portion is at least partially located below the waste outlet.

52. The system of claim 50, further comprising a discrete spacing fluid reservoir configured to contain a spacing fluid for input into the sample reader via a spacing fluid inlet, and a discrete waste reservoir configured to receive waste from the sample reader via a waste outlet, wherein the sample reader is configured to interchangeably use a combined spacing fluid and waste reservoir or a discrete spacing fluid reservoir and a discrete waste reservoir while analyzing a sample.

53. A system according to claim 52, wherein the separation between the openings in the combination reservoir is the same as the separation between the openings in the discrete spacer fluid reservoir and the discrete waste reservoir when the discrete reservoirs are properly positioned for use with a sample reader.

54. The system of claim 50, wherein each opening is connected to the common volume by a neck, and wherein the neck associated with one opening is wider than the neck associated with another opening.

55. The system of claim 50, wherein the accessible fluid depth directly beneath one opening is different than the accessible fluid depth directly beneath another opening.

56. The system of claim 55, wherein the spacer fluid is optionally denser than the sample, wherein the spacer fluid inlet accesses the reservoir through an opening associated with a greater accessible fluid depth, and the waste outlet accesses the reservoir through an opening associated with a lesser accessible fluid depth.

57. The system of claim 55, wherein the combined reservoir has a bottom configured to rest on a surface when the reservoir is in use, wherein each opening is the same distance from the bottom.

58. The system of claim 47, wherein the combined reservoir has a single opening and wherein a trap fluid configured to contain additional spacer fluid and waste is inserted between the reservoir and the sample reader.

59. The system of claim 58, wherein the sample reader receives spacer fluid from the trap and returns waste to the trap.

60. The system of claim 59, wherein the spacer fluid and waste material are exchanged with fluid in the reservoir such that waste material moves from the trap to the reservoir and the spacer fluid moves from the reservoir to the trap.

61. The system of claim 58, wherein the sample reader further has an onboard reservoir for holding a spacing fluid received from the trap, and at least one of a waste tank for holding waste material before it is returned to the tank.

62. The system of claim 47, wherein the partitions are droplets, the system further comprising a droplet generator configured to divide the sample into droplets before the sample is analyzed by a sample reader.

63. The system of claim 62, further comprising a thermal cycler configured to thermally cycle the droplets after the droplet generator generates the droplets and before the sample reader analyzes the droplets.

64. The system of claim 47, wherein the volume of the combined reservoir is less than or equal to about 1500 mL.

65. The system of claim 64, wherein the volume of the combined reservoir is about 1000 mL.

66. The system of claim 47, wherein the spacer fluid has a greater density than the sample, wherein the spacer fluid inlet is positioned toward a bottom of the combination reservoir.

67. The system of claim 47, the sample having a density greater than a density of the spacer fluid, wherein the spacer fluid inlet is positioned toward a top of the combination reservoir.

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