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        Reliable medical centrifugation has historically required the use of expensive, bulky, and electrically dependent commercial equipment, which is often not available in resource-limited settings. Although several portable, inexpensive, non-motorized centrifuges have been described, these solutions are primarily intended for diagnostic applications requiring relatively small volumes of sedimentation. In addition, the design of these devices often requires the use of special materials and tools that are not normally available in underserved areas. Here we describe the design, assembly, and experimental validation of the CentREUSE, an ultra-low cost, human-operated, portable waste-based centrifuge for therapeutic applications. CentREUSE exhibits an average centrifugal force of 10.5 relative centrifugal force (RCF) ± 1.3. Settling of 1.0 ml vitreous suspension of triamcinolone after 3 minutes of centrifugation in CentREUSE was comparable to that after 12 hours of gravity-mediated sedimentation (0.41 ml ± 0.04 vs 0.38 ml ± 0.03, p = 0.14). Sediment thickening after CentREUSE centrifugation for 5 and 10 minutes compared to that observed after centrifugation at 10 RCF (0.31 ml ± 0.02 vs. 0.32 ml ± 0.03, p = 0.20) and 50 RCF (0.20 ml) for 5 minutes using commercial equipment Similar ± 0.02 vs. 0.19 ml ± 0.01, p = 0.15). The templates and building instructions for CentREUSE are included in this open source post.
        Centrifugation is an important step in many diagnostic tests and therapeutic interventions1,2,3,4. However, achieving adequate centrifugation has historically required the use of expensive, bulky, and electrically dependent commercial equipment, which is often not available in resource-limited settings2,4. In 2017, Prakash’s group introduced a small paper-based manual centrifuge (called a “paper puffer”) made from prefabricated materials at a cost of $0.20 ($)2. Since then, paper fugue has been deployed in resource-limited settings for low-volume diagnostic applications (e.g. density-based separation of blood components in capillary tubes to detect malaria parasites), thus demonstrating a super-cheap portable human-powered instrument. centrifuge 2 . Since then, several other compact, inexpensive, non-motorized centrifugation devices have been described4,5,6,7,8,9,10. However, most of these solutions, like paper fumes, are intended for diagnostic purposes requiring relatively small sedimentation volumes and therefore cannot be used to centrifuge large samples. In addition, the assembly of these solutions often requires the use of special materials and tools that are often not available in underserved areas4,5,6,7,8,9,10.
        Here we describe the design, assembly, and experimental validation of a centrifuge (called the CentREUSE) constructed from conventional paper fugue waste for therapeutic applications that typically require high sedimentation volumes. Case 1, 3 As a proof of concept, we tested the device with a real ophthalmic intervention: precipitation of a suspension of triamcinolone in acetone (TA) for subsequent injection of a bolus drug into the vitreous body of the eye. Although centrifugation for TA concentration is a recognized low-cost intervention for the long-term treatment of various eye conditions, the need for commercially available centrifuges during drug formulation is a major barrier to the use of this therapy in resource-limited settings1,2,3. compared with results obtained with conventional commercial centrifuges. Templates and instructions for building CentREUSE are included in this open source posting in the “More Information” section.
        CentREUSE can be built almost entirely from scrap. Both copies of the semi-circular template (Supplementary Figure S1) were printed on standard US carbon letter paper (215.9 mm × 279.4 mm). The attached two semi-circular templates define three key design features of the CentREUSE device, including (1) the outer rim of the 247mm spinning disk, (2) is designed to accommodate a 1.0ml syringe (with cap and amputated plunger). grooves in the shank) and (3) two marks indicating where to punch holes so that the rope can pass through the disk.
        Adhere (e.g. with all-purpose adhesive or tape) the template to the corrugated board (minimum size: 247 mm × 247 mm) (Supplementary Figure S2a). Standard “A” corrugated board (4.8 mm thick) was used in this study, but corrugated board of similar thickness could be used, such as corrugated board from discarded shipping boxes. Using a sharp tool (such as a blade or scissors), cut the cardboard along the edge of the outer disc outlined on the template (Supplementary Figure S2b). Then, using a narrow, sharp tool (such as the tip of a ballpoint pen), create two full-thickness perforations with a radius of 8.5 mm according to the marks traced on the template (Supplementary Figure S2c). Two slots for 1.0 ml syringes are then cut from the template and the underlying surface layer of cardboard using a pointed tool such as a razor blade; care must be taken not to damage the underlying corrugated layer or the remaining surface layer (Supplementary Figure S2d, e) . Then, thread a piece of string (e.g. 3mm cooking cotton cord or any thread of similar thickness and elasticity) through the two holes and tie a loop around each side of a disc about 30cm long (Supplementary Fig. S2f).
        Fill two 1.0 ml syringes with approximately equal volumes (eg 1.0 ml of TA suspension) and cap. The syringe plunger rod was then cut off at the level of the barrel flange (Supplementary Figure S2g, h). The cylinder flange is then covered with a layer of tape to prevent the ejection of the truncated piston during use of the equipment. Each 1.0 ml syringe was then placed in the syringe well with the cap facing the center of the disc (Supplementary Figure S2i). Each syringe was then attached to at least the disc with adhesive tape (Supplementary Figure S2j). Finally, complete the assembly of the centrifuge by placing two pens (such as pencils or similar sturdy stick-shaped tools) at each end of the string inside the loop (Figure 1).
        Instructions for using the CentREUSE are similar to those for traditional spinning toys. The rotation is started by holding a handle in each hand. Slight slack in the strings causes the disc to rock forward or backward, causing the disc to rotate forward or backward respectively. This is done several times in a slow, controlled manner so that the strings curl up. Then stop the movement. As the strings begin to unwind, the handle is pulled hard until the strings are taut, causing the disc to spin. As soon as the string is completely unwound and begins to rewind, the handle should be slowly relaxed. As the rope begins to unwind again, apply the same series of motions to keep the device spinning (video S1).
        For applications requiring sedimentation of a suspension by centrifugation, the device was continuously rotated until satisfactory granulation was achieved (Supplementary Figure S3a,b). Complex particles will form at the plunger end of the syringe barrel and the supernatant will concentrate towards the tip of the syringe. The supernatant was then drained by removing the tape covering the barrel flange and introducing a second plunger to slowly push the native plunger towards the syringe tip, stopping when it reached the compound sediment (Supplementary Figure S3c,d).
        To determine the rotation speed, the CentREUSE device, equipped with two 1.0 ml syringes filled with water, was recorded with a high-speed video camera (240 frames per second) for 1 min after reaching a steady state of oscillation. Markers near the edge of the spinning disk were tracked manually using frame-by-frame analysis of the recordings to determine the number of revolutions per minute (rpm) (Figures 2a-d). Repeat n = 10 attempts. The relative centrifugal force (RCF) at the midpoint of the syringe barrel is then calculated using the following formula:
        Rotational speed quantification with CentREUSE. (A–D) Sequential representative images showing the time (minutes: seconds. milliseconds) to complete device rotation. Arrows indicate trace markers. (E) RPM quantification using CentREUSE. The lines represent the mean (red) ± standard deviation (black). The scores represent individual 1-minute trials (n = 10).
        A 1.0 ml syringe containing TA suspension for injection (40 mg/ml, Amneal Pharmaceuticals, Bridgewater, NJ, USA) was centrifuged for 3, 5 and 10 minutes using CentREUSE. Sedimentation using this technique was compared with that achieved after centrifugation at 10, 20, and 50 RCF using an A-4-62 rotor for 5 min on an Eppendorf 5810R benchtop centrifuge (Hamburg, Germany). The amount of precipitation was also compared with the amount of precipitation obtained using gravity-dependent precipitation at various time points from 0 to 720 minutes. A total of n = 9 independent repetitions were performed for each procedure.
        All statistical analyzes were performed using Prism 9.0 software (GraphPad, San Diego, USA). Values ​​are presented as mean ± standard deviation (SD) unless otherwise noted. Group means were compared using a two-tailed Welch-corrected t-test. Alpha is defined as 0.05. For gravity-dependent subsidence, a single-phase exponential decay model was fitted using least-squares regression, treating repeated y values ​​for a given x value as a single point.
        where x is the time in minutes. y – sediment volume. y0 is the value of y when x is zero. The plateau is the y value for infinite minutes. K is the rate constant, expressed as the reciprocal of minutes.
        The CentREUSE device demonstrated reliable, controlled non-linear oscillations using two standard 1.0 ml syringes filled with 1.0 ml of water each (video S1). In n = 10 trials (1 minute each), CentREUSE had an average rotational speed of 359.4 rpm ± 21.63 (range = 337-398), resulting in a calculated average centrifugal force of 10.5 RCF ± 1, 3 (range = 9.2–12.8). (Figure 2a-e).
        Several methods for pelleting TA suspensions in 1.0 ml syringes were evaluated and compared with CentREUSE centrifugation. After 12 hours of gravity-dependent settling, the sediment volume reached 0.38 ml ± 0.03 (Supplementary Fig. S4a,b). Gravity-dependent TA deposition is consistent with a single-phase exponential decay model (corrected by R2 = 0.8582), resulting in an estimated plateau of 0.3804 mL (95% confidence interval: 0.3578 to 0.4025) (Supplementary Figure S4c). CentREUSE produced an average sediment volume of 0.41 ml ± 0.04 at 3 minutes, which was similar to the mean value of 0.38 ml ± 0.03 observed for gravity-dependent sedimentation at 12 hours (p = 0.14) (Fig. 3a, d, h). CentREUSE gave a significantly more compact volume of 0.31 ml ± 0.02 at 5 minutes compared to the mean of 0.38 ml ± 0.03 observed for gravity-based sedimentation at 12 hours (p = 0.0001) ( Fig. 3b, d, h).
        Comparison of TA pellet density achieved by CentREUSE centrifugation with gravity settling versus standard industrial centrifugation (A–C). Representative images of precipitated TA suspensions in 1.0 ml syringes after 3 min (A), 5 min (B), and 10 min (C) of CentREUSE use. (D) Representative images of deposited TA after 12 h of gravity settling. (EG) Representative images of precipitated TA after standard commercial centrifugation at 10 RCF (E), 20 RCF (F), and 50 RCF (G) for 5 min. (H) Sediment volume was quantified using CentREUSE (3, 5, and 10 min), gravity-mediated sedimentation (12 h), and standard industrial centrifugation at 5 min (10, 20, and 50 RCF). The lines represent the mean (red) ± standard deviation (black). The dots represent independent repeats (n = 9 for each condition).
        CentREUSE produced a mean volume of 0.31 ml ± 0.02 after 5 minutes, which is similar to the mean of 0.32 ml ± 0.03 observed in a standard commercial centrifuge at 10 RCF for 5 minutes (p = 0.20), and slightly lower than the mean volume obtained with 20 RCF was observed at 0.28 ml ± 0.03 for 5 minutes (p = 0.03) (Fig. 3b, e, f, h). CentREUSE produced a mean volume of 0.20 ml ± 0.02 at 10 minutes, which was just as compact (p = 0.15) compared to a mean volume of 0.19 ml ± 0.01 at 5 minutes observed with a commercial centrifuge at 50 RCF (Fig. 3c, g, h). .
        Here we describe the design, assembly, and experimental verification of an ultra-low-cost, portable, human-operated, paper-based centrifuge made from conventional therapeutic waste. The design is largely based on the paper-based centrifuge (referred to as “paper fugue”) introduced by Prakash’s group in 2017 for diagnostic applications. Given that centrifugation has historically required the use of expensive, bulky, and electrically dependent commercial equipment, Prakash’s centrifuge provides an elegant solution to the problem of insecure access to centrifugation in resource-limited settings2,4. Since then, paperfuge has shown practical utility in several low-volume diagnostic applications, such as density-based blood fractionation for malaria detection. However, to the best of our knowledge, similar ultra-cheap paper-based centrifuge devices have not been used for therapeutic purposes, conditions that typically require larger volume sedimentation.
        With this in mind, CentREUSE’s goal is to expand the use of paper centrifugation in therapeutic interventions. This was achieved by making several modifications to the design of the Prakash reveal. Notably, to increase the length of two standard 1.0 ml syringes, CentREUSE contains a larger disk (radius = 123.5 mm) than the largest Prakash paper wringer tested (radius = 85 mm). In addition, to support the extra weight of a 1.0 ml syringe filled with liquid, CentREUSE uses corrugated cardboard instead of cardboard. Together, these modifications allow centrifugation of larger volumes than those tested in the Prakash paper cleaner (i.e. two 1.0 ml syringes with capillaries) while still relying on similar components: filament and paper-based material. Notably, several other inexpensive human-powered centrifuges have been described for diagnostic purposes4,5,6,7,8,9,10. These include spinners, salad beaters, egg beaters, and hand torches for rotating devices5, 6, 7, 8, 9. However, most of these devices are not designed to handle volumes up to 1.0 ml and consist of materials that are often more expensive and inaccessible than those used in paper centrifuges2,4,5,6,7,8,9,10. . In fact, discarded paper materials are often found everywhere; for example, in the United States, paper and paperboard account for over 20% of municipal solid waste, providing a plentiful, inexpensive, or even free source for building paper centrifuges. e.g. CentREUSE11. Also, compared to several other low cost solutions published, CentREUSE does not require specialized hardware (such as 3D printing hardware and software, laser cutting hardware and software, etc.) to create, making the hardware more resource intensive. . These people are in the environment 4, 8, 9, 10.
        As proof of the practical usefulness of our paper centrifuge for therapeutic purposes, we demonstrate the rapid and reliable settling of triamcinolone suspension in acetone (TA) for vitreous bolus injection—an established low-cost intervention for the long-term treatment of various ocular diseases1,3. Settling results after 3 minutes with CentREUSE were comparable to results after 12 hours of gravity-mediated settling. In addition, CentREUSE results after centrifugation for 5 and 10 minutes exceeded results that would be obtained by gravity and were similar to those observed after industrial centrifugation at 10 and 50 RCF for 5 minutes, respectively. Notably, in our experience, CentREUSE produces a sharper and smoother sediment-supernatant interface than other methods tested; this is desirable as it allows for a more accurate assessment of the dose of the administered drug, and it is easier to remove the supernatant with minimal loss of particle volume.
        The choice of this application as a proof of concept was driven by the ongoing need to improve access to long-acting intravitreal steroids in resource-limited settings. Intravitreal steroids are widely used to treat a variety of eye conditions, including diabetic macular edema, age-related macular degeneration, retinal vascular occlusion, uveitis, radiation retinopathy, and cystic macular edema3,12. Of the steroids available for intravitreal administration, TA remains the most commonly used worldwide12. Although preparations without TA preservatives (PF-TA) are available (eg, Triesence [40 mg/mL, Alcon, Fort Worth, USA]), preparations with benzyl alcohol preservatives (eg, Kenalog-40 [40 mg/mL, Bristol-Myers Squibb, New York, USA]) remains the most popular3,12. It should be noted that the latter group of drugs is approved by the US Food and Drug Administration (FDA) for intramuscular and intraarticular use only, so intraocular administration is considered unregistered 3, 12 . Although the injectable dose of intravitreal TA varies according to indication and technique, the most commonly reported dose is 4.0 mg (i.e. injection volume of 0.1 ml from a 40 mg/ml solution), which usually gives a treatment duration of approximately 3 months Effects 1, 12, 13, 14, 15.
        To prolong the action of intravitreal steroids in chronic, severe or recurrent eye diseases, several long-acting implantable or injectable steroid devices have been introduced, including dexamethasone 0.7 mg (Ozurdex, Allergan, Dublin, Ireland), Relax fluoride acetonide 0.59 mg (Retisert, Bausch and Lomb, Laval, Canada) and fluocinolone acetonide 0.19 mg (Iluvien, Alimera Sciences, Alpharetta, Georgia, USA)3,12. However, these devices have several potential drawbacks. In the United States, each device is only approved for a few indications, limiting insurance coverage. In addition, some devices require surgical implantation and may cause unique complications such as device migration into the anterior chamber3,12. In addition, these devices tend to be less readily available and much more expensive than TA3,12; at current US prices, Kenalog-40 costs about $20 per 1.0 ml of suspension, while Ozurdex, Retisert, and Iluvien explants. The entrance fee is about $1400. , $20,000 and $9,200 respectively. Together, these factors limit access to these devices for people in resource-constrained settings.
        Attempts have been made to prolong the effect of intravitreal TA1,3,16,17 due to its lower cost, more generous reimbursement, and greater availability. Given its low water solubility, TA remains in the eye as a depot, allowing gradual and relatively constant drug diffusion, so the effect is expected to last longer with larger depots1,3. To this end, several methods have been developed to concentrate the TA suspension prior to injection into the vitreous. Although methods based on passive (ie gravity dependent) settling or microfiltration have been described, these methods are relatively time consuming and give variable results15,16,17. On the contrary, previous studies have shown that TA can be rapidly and reliably concentrated (and thus prolonged action) by centrifugation-assisted precipitation1,3. In conclusion, the convenience, low cost, duration, and efficacy of centrifugally concentrated TA make this intervention an attractive option for patients in resource-limited settings. However, lack of access to reliable centrifugation can be a major barrier to implementing this intervention; By addressing this issue, CentREUSE can help increase the availability of long-term steroid therapy for patients in resource-limited settings.
        There are some limitations in our study, including those related to functionality native to the CentREUSE appliance. The device is a non-linear, non-conservative oscillator that relies on human input and therefore cannot provide an accurate and constant rotation rate during use; rotation speed depends on several variables, such as user influence on the level of device ownership, specific materials used in the assembly of equipment, and the quality of the connections being spun. This is different from commercial equipment where rotational speed can be applied consistently and accurately. In addition, the speed achieved by CentREUSE can be considered relatively modest compared to the speed achieved by other centrifuge devices2. Fortunately, the speed (and associated centrifugal force) generated by our device was sufficient to test the concept detailed in our study (i.e., TA deposition). The rotation speed can be increased by lightening the mass of the central disk 2; this can be achieved by using a lighter material (such as thinner cardboard) if it is strong enough to hold two syringes filled with liquid. In our case, the decision to use standard “A” slotted cardboard (4.8 mm thick) was deliberate, as this material is often found in shipping boxes and is therefore easily found as a recyclable material. The rotation speed can also be increased by reducing the radius of the central disk 2 . However, the radius of our platform was deliberately made relatively large to accommodate a 1.0 ml syringe. If the user is interested in centrifuging shorter vessels, the radius can be reduced—a change that predictably results in higher rotational speeds (and possibly higher centrifugal forces).
        In addition, we have not carefully evaluated the impact of operator fatigue on equipment functionality. Interestingly, several members of our group were able to use the device for 15 minutes without noticeable fatigue. A potential solution to operator fatigue when longer centrifuges are required is to rotate two or more users (if possible). In addition, we did not critically evaluate the durability of the device, in part because the components of the device (such as cardboard and cord) could be easily replaced at little or no cost in the event of wear or damage. Interestingly, during our pilot test, we used one device for a total of over 200 minutes. After this period, the only noticeable but minor sign of wear is perforation along the threads.
        Another limitation of our study is that we did not specifically measure the mass or density of deposited TA, achievable with the CentREUSE device and other methods; instead, our experimental verification of this device was based on the measurement of sediment density (in ml). indirect measure of density. In addition, we have not tested CentREUSE Concentrated TA on patients, however, since our device produced TA pellets similar to those produced using a commercial centrifuge, we assumed that CentREUSE Concentrated TA would be as effective and safe as previously used. in literature. reported for conventional centrifuge devices1,3. Additional studies quantifying the actual amount of TA administered after CentREUSE fortification may help further evaluate the actual usefulness of our device in this application.
        To our knowledge, CentREUSE, a device that can be easily constructed from readily available waste, is the first human-powered, portable, ultra-low-cost paper centrifuge to be used in a therapeutic setting. In addition to being able to centrifuge relatively large volumes, CentREUSE does not require the use of specialized materials and construction tools compared to other low cost centrifuges published. The demonstrated efficacy of CentREUSE in rapid and reliable TA precipitation may help improve long-term intravitreal steroid availability in people in resource-limited settings, which may help treat a variety of ocular conditions. In addition, the benefits of our portable human-powered centrifuges predictably extend to resource-rich locations such as large tertiary and quaternary health centers in developed countries. Under these conditions, the availability of centrifuging devices may continue to be limited to clinical and research laboratories, with the risk of contaminating syringes with human body fluids, animal products, and other hazardous substances. In addition, these laboratories are often located far from the point of care for patients. This, in turn, can be a logistical hurdle for healthcare providers who need quick access to centrifugation; deploying CentREUSE can serve as a practical way to prepare therapeutic interventions in the short term without seriously disrupting patient care.
        Therefore, to make it easier for everyone to prepare for therapeutic interventions that require centrifugation, a template and instructions for creating CentREUSE are included in this open source publication under the Additional Information section. We encourage readers to redesign CentREUSE as needed.
       Data supporting the results of this study are available from the respective SM author upon reasonable request.
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       SM is supported in part by a gift to the Mukai Foundation, Massachusetts Eye and Ear Hospital, Boston, Massachusetts, USA.
       Department of Ophthalmology, Harvard Medical School, Massachusetts Eye and Ear, 243 Charles St, Boston, Massachusetts, 02114, USA