Use of U-500 insulin in the treatment of severe insulin resistance

Background: Glycemic control is essential in the management of diabetes. However, many patients with diabetes are not achieving therapeutic targets, partly because they are receiving insufficient doses of insulin. This is particularly problematic in patients with severe insulin resistance, defined as insulin requirement >200 units/kg per day (>3 units/kg per day for pediatric patients). It is difficult to use U-100 forms of insulin at doses >200 units/kg per day because of the volume of insulin being administered subcutaneously. U-500, a concentrated form of insulin, may be useful in the treatment of these patients.Objective: Current practice regarding the use of U-500 insulin has been published elsewhere. This article presents an updated algorithm for the administration and dosing of U-500 insulin, based on clinical experience with severe forms of insulin resistance. Guidelines are provided for dose escalation of U-500 insulin.Methods: We reviewed the results of treatment with U-500 insulin in patients with severe insulin resistance. We analyzed the results, updated a pre-existing algorithm, provided additional practical information on the administration and dosing of U-500 insulin, and compared the cost of U-500 with that of U-100 insulin.Results: To date, we have treated 56 patients (age range, 9–54 years) with severe insulin resistance using U-500 insulin. Doses ranged from 1.5 to 566 units/kg per day. Based on the pharmacodynamic properties of U-500 insulin, this concentrated form must be administered and dosed differently than regular U-100 insulin. U-500 insulin cost more than U-100 insulin on a per-milliliter basis, but cost less in the end because of the lower volumes of insulin required and fewer syringes and pump cartridges needed to administer U-500 insulin.Conclusions: In our experience, U-500 insulin is a useful tool in the management of patients with severe insulin resistance. U-500 insulin alleviates the volume-related problems associated with U-100 insulin, making treatment with higher doses of insulin (≥200 units per day) more effective with U-500 insulin than with U-100 insulin.     

Effect of Total Daily Dose on Efficacy,Dosing, and Safety of 2 Dose Titration Regimens of Human Regular U-500 Insulin in Severely Insulin-Resistant Patients with Type 2 Diabetes

Objective: To examine the influence of baseline U-100 insulin total daily dose (TDD) on clinical outcomes in severely insulin-resistant patients with inadequately controlled type 2 diabetes treated with human regular U-500 insulin (U-500R) from the perspective of current dosing recommendations.Methods: Data from a recent prospective, randomized trial comparing thrice-daily (TID) and twice-daily (BID) U-500R in 325 patients transitioned from highdose/high-volume U-100 insulin were analyzed across baseline U-100 TDD units and units/kg subgroups (≤300 units [n = 224, 68.9%] and >300 units [n = 101, 31.1%]; ≤2 units/kg [n = 96, 29.5%] and >2 units/kg [n = 229, 70.5%]). Subgroup effects on treatment differences were evaluated, and outcomes between treatment-pooled subgroups were compared.Results: At 24 weeks, significant reductions in glycated hemoglobin (HbA1c) were observed for all subgroups (range: -1.01% to -1.38%, P<.05). Within-subgroup treatment effects were similar with no treatment-by-subgroup interactions; however, a greater reduction was noted in the >300-units subgroup (P = .04). No TID/BID differences within subgroups or treatment-by-subgroup interactions were observed for TDD or weight increase from baseline. Overall hypoglycemia rates were similar between treatments (within subgroups) and showed no interactions. However, rates were higher in the >300-units subgroup for severe hypoglycemia (P = .04) and in both higher-dose subgroups for documented symptomatic hypoglycemia ≤70 mg/dL (P<.001, units; P = .001, units/kg).Conclusion: Both TID and BID U-500R were efficacious and safe across TDD subgroups, though higher hypoglycemia rates were observed in higher-dose, treatment-pooled subgroups. U-500R dosing recommendations have been updated accordingly.Abbreviations:AE = adverse eventBID = twice dailyHbA1c = glycated hemoglobinQID = 4 times dailyRCT = randomized clinical trialT2D = type 2 diabetesTDD = total daily doseTID = thrice dailyU-500R = human regular U-500 insulin     

High-dose Insulin Therapy: is it Time for U-500 Insulin?

ObjectiveTo provide an overview of U-500 regular insulin action, review published clinical studies with U- 500 regular insulin, and offer guidance to practicing endocrinologists for identifying patients for whom U-500 regular insulin may be appropriate.MethodsThis review has been produced through a synthesis of relevant published literature compiled via a literature search (MEDLINE search of the English-language literature published between January 1969, and July 2008, related to U-500, insulin resistance, concentrated insulin, high-dose insulin, insulin pharmacokinetics, and diabetes management) and the authors’ collective clinical experience.ResultsThe obesity epidemic is contributing to an increase in the prevalence of type 2 diabetes, as well as to increasing insulin requirements in insulin-treated patients. Many of these patients exhibit severe insulin resistance, manifested by daily insulin requirements of 200 units or greater or more than 2 units/kg. Delivering an appropriate insulin volume to these patients can be difficult and inconvenient and may be best accomplished with U-500 regular insulin by multiple daily injections or with continuous subcutaneous insulin infusion, rather than with standard U-100 insulin. Implementation of U-500 regular insulin in patients previously on other insulin formulations is described with a treatment algorithm covering dosage requirements ranging from 150 to more than 600 units per day on the basis of the authors’ experience.ConclusionRegimen conversion of appropriately selected patients from high-dose, U-100 insulin to U-500 regular insulin therapy on the basis of the recommendations presented in this article may potentially result in improved glycemic control and lower cost. (Endocr Pract. 2009;15:71-79)     

High antigenicity of intraperitoneal insulin infusion via implantable devices: preliminary rat studies.

Intraperitoneal insulin infusion of Genapol stabilized insulin via implantable devices significantly improves diabetes control and hypoglycemia frequency in type 1 diabetes while it increases insulin antibody levels. Causes for this particular antigenicity remain unknown. The role of insulin modifications occurring in the reservoir on the antigenicity observed was assessed by comparing the antigenicities of the insulin coming from the vial or from the pump reservoir. Rats were injected intraperitoneally with insulin sampled either from a vial (group 1) or from a pump reservoir during a refill of a clinical trial (group 2). Two control groups, one without insulin, the second one receiving a mixture of silicone and insulin were also studied. Human insulin antibody levels were assessed by RIA 10 days after 4 weekly immunizations. AIA levels were higher in group 1 compared to group 2 (P = 0.003 for the first experiment, P = 0.04 in the second experiment). The increased antigenicity of the insulin sampled from the implanted pump might be due to the insulin modifications occurring during the storage in the device. Insulin aggregates could be involved in this antigenicity since they are known to be antigenic and their concentration was shown to be related to the amplitude of the antigenic response.     

Two Treatment Approaches for Human Regular U-500 Insulin in Patients With Type 2 Diabetes not Achieving Adequate Glycemic Control on High-Dose U-100 Insulin Therapy With or Without Oral Agents: A Randomized,Titration-To-Target Clinical Trial

Objective: To compare the efficacy and safety of 2 dosing regimens for human regular U-500 insulin (U-500R, 500 units/mL) replacing high-dose U-100 insulins with or without oral antihyperglycemic drugs in patients with inadequately controlled type 2 diabetes (T2D).Methods: We conducted a 24-week, open-label, parallel trial in 325 patients (demographics [means]: age, 55.4 years; diabetes duration, 15.2 years; body mass index, 41.9 kg/m²; glycated hemoglobin [HbA_1c], 8.7%; U-100 insulin dose, 287.5 units administered in 5 injections/day [median; range, 2 to 10]). Patients were randomized to thrice-daily (TID, n = 162) or twice-daily (BID, n = 163) U-500R after a 4-week lead-in period. The primary outcome was HbA_1c change from baseline.Results: After 24 weeks, both treatments demonstrated significant HbA_1c reductions (TID, -1.12%; BID, -1.22%; both, P<.001) and clinical equivalence (difference, -0.10%; 95% confidence interval, -0.33 to 0.12%; noninferiority margin, 0.4%). Comparable increases in total daily U-500R dose (TID, 242.7 to 343.1 units; BID, 249.0 to 335.0 units) were observed. Incidence and rate of documented symptomatic hypoglycemia (≤70 mg/dL) were lower for TID versus BID (P = .003 and P = .02, respectively); severe hypoglycemia was similar between treatments. Weight gain was similar in both groups (TID, 5.4 kg; BID, 4.9 kg).Conclusion: Initiation and titration of U-500R using either algorithm (TID or BID) improves glycemic control effectively and safely with fewer injections in patients with T2D treated with high-dose/high-volume U-100 insulin. These results provide clinicians with a practical framework for using U-500R in severely insulin-resistant patients with suboptimally controlled T2D.Abbreviations: BID = twice daily FAS = full analysis set HbA_1c = glycated hemoglobin PG = plasma glucose SMPG = self-monitored plasma glucose T2D = type 2 diabetes TDD = total daily dose TID = thrice daily U-500R = human regular U-500 insulin     

Evaluation of Total Daily Dose and Glycemic Control for Patients Taking U-500 Regular Insulin Admitted to the Hospital

Objective: Patients using U-500 regular insulin are severely insulin resistant, requiring high doses of insulin. It has been observed that a patient's insulin requirements may dramatically decrease during hospitalization. This study sought to systematically investigate this phenomenon.Methods: We performed a retrospective chart review of patients with U-500 insulin outpatient regimens who were admitted to the San Antonio Military Medical Center over a 5-year period. Each patient's outpatient total daily dose (TDD) of insulin was compared to the average inpatient TDD. The outpatient estimated average glucose (eAG) was calculated from the glycated hemoglobin (HbA1c) and compared to the average inpatient glucose.Results: There were 27 patients with a total of 62 separate admissions. The average age was 64.4 years, with a mean body mass index of 38.9 kg/m² and eAG of 203 mg/dL (HbA1c, 8.7%, 71.6 mmol/mol). All patients were converted from U-500 to U-100 upon admission. The average inpatient TDD of insulin was 91 units, versus 337 units as outpatients (P<.001). Overall, 89% of patients received ≤50% of their outpatient TDD. The average inpatient glucose was slightly higher than the outpatient eAG, 234 mg/dL versus 203 mg/dL (P = .003).Conclusion: U-500 insulin is prone to errors in the hospital setting, so conversion to U-100 insulin is a preferred option. Despite a significant reduction in insulin TDD, these patients had clinically similar glucose levels. Therefore, patients taking U-500 insulin as an outpatient can be converted to a U-100 basal-bolus regimen with at least a 50% reduction of their outpatient TDD.Abbreviations:BG = blood glucoseeAG = estimated average glucoseHbA1c = glycated hemoglobinNPO = nil per osSPSS = Statistical Package for the Social SciencesTDD = total daily dose     

U-73122 does not specifically inhibit phospholipase C in rat pancreatic islets and insulin-secreting β-cell lines

Phospholipase C is activated in insulin secretion by islets of Langerhans and insulin-secreting β-cells such as RINm5F and β-TC3. We have examined the effects of the aminosteroid U-73122, a phospholipase C inhibitor, on insulin secretion and phospholipase C activation. U-73122 slightly inhibited glucose-induced insulin secretion from islets, but this effect was not specific since the structural “inactive” analogue U-73343 also inhibited insulin secretion. Likewise, in RINm5F cells, U-73122 did not inhibit glyceraldehyde-induced insulin secretion. Phospholipase C activity was assessed as the accumulation of inositol-1,4,5-trisphosphate (Ins(1,4,5)P₃) measured with a competitive binding assay: U-73122 failed to inhibit glucose-induced increase in Ins(1,4,5)P₃. Similarly, when the effects of U-73122 and U-73343 were measured on [³H]phosphatidylinositol hydrolysis of islets, both compounds caused a slight, non-specific inhibition of phospholipase C activity. These observations suggest that U-73122 does not specifically inhibit phospholipase C in insulin-secreting cells.     

Theoretical analysis of the effect of convective flow on solute transport and insulin release in a hollow fiber bioartificial pancreas

The bioartificial pancreas, in which transplanted pancreatic tissue or isolated cells are cultured on a hollow fiber membrane, is an attractive approach to restore physiologic insulin delivery in the treatment of diabetes. Insulin response in prototype devices has been unacceptable due to the large mass transport limitations associated with the membrane and the surrounding shell region. Although available theoretical analyses provide some insight into the combined effects of transport and reaction in the bioartificial pancreas, they cannot quantitatively account for the effects of convective recirculation flow, complex intrinsic insulin secretory kinetics, and non-uniform distribution of pancreatic cells. We have developed a detailed model for glucose and insulin transport and insulin secretion in the hollow fiber bioartificial pancreas based on the solution of the mass and momentum conservation equations describing flow and transport in the lumen, matrix, and shell. Model predictions are in good agreement with literature data obtained in a hollow fiber device with minimal radial convective flow. Although no quantitative data are available for a device with significant radial convection, model simulations demonstrate that convective recirculation flow can dramatically improve insulin response, allowing the device to accurately capture the bi-phasic insulin secretion characteristic of the normal physiologic response. Results provide fundamental insights into the coupling between kinetics and transport in the hollow fiber system and a rational basis for the design of clinical devices.     

An open-loop insulin delivery device for the control of experimental diabetes.

A new insulin delivery device has been developed and tested. It includes a reservoir, a pump, and a power pack. The reservoir holds 75 ml and is coupled to a precision peristaltic pump whose delivery can be set to any one of 128 different flow rates from 0 to 80 microliter/min (+/- 1.6% over 10 months) using the flow rate controller included in the battery power pack. The system weighs 525 g, consuming 50 mW at the maximum pumping rate, proportionately less at lower rates. Ten pumps have undergone bench tests for 30 days. One has been subjected to an extended life test of 16 months without change of tubing while seven complete systems have been used on dogs to demonstrate their capability for precise long-term (up to 16 months) intravenous insulin therapy. With this system, experimental diabetes has been controlled in 7 dogs for periods now extending beyond 16 months. This device now qualifies for-long term studies on hospitalized patients with diabetes mellitus.     

Clinical Efficacy and Patient Satisfaction with U-500 Insulin Pump Therapy in Patients with Type 2 Diabetes

ObjectiveTo determine the safety and efficacy of U-500 regular insulin in pump therapy and to assess satisfaction with U-500 insulin pump therapy in patients with type 2 diabetes and severe insulin resistance.MethodsWe performed a retrospective review of medical records of 6 patients with type 2 diabetes and insulin resistance who had been using U-500 insulin pump therapy for at least 6 months. In addition, we conducted a telephone follow-up patient satisfaction survey.ResultsThe mean age of the patients was 57 ± 6 years. Of the 6 patients (3 men and 3 women), 4 were white, 1 was black, and 1 was Asian. The mean hemoglobin A1c value before continuous subcutaneous insulin infusion therapy with U-500 regular insulin was 9.1% ± 1.8%, and the mean U-100 insulin dose required was 391 ± 91 U/day. In comparison, the mean U-500 insulin dose required at 6 months was 59.2 ± 13.6 U/day, which is equivalent to 296 ± 68 U/day of U-100 insulin (P = 0.04), and the mean hemoglobin A1c value at 6 months after treatment with the insulin pump using U-500 regular insulin was 6.9% ± 0.9% (P = 0.03). In addition, our study patients lost a mean of 6.1 lb during the 6-month period, and no patient had clinically significant episodes of hypoglycemia. The majority of the patients reported a higher satisfaction with the U-500 insulin pump therapy in comparison with conventional insulin treatment.ConclusionU-500 insulin pump therapy is a novel alternative for patients with type 2 diabetes and severe insulin resistance who have not met glycemic control goals with use of standard intensive insulin regimens. (Endocr Pract. 2007;13:721-725)     

Down-regulation and recycling of insulin receptors. Effect of monensin on IM-9 lymphocytes and U-937 monocyte-like cells

Receptor down-regulation is the result of various cellular processes including receptor internalization, new synthesis, and recycling. Monensin, a monocarboxylic acid ionophore, has been used to characterize the role of recycling in the metabolism of insulin receptors on two cultured human cell lines, U-937 and IM-9, which have different rates of internalization. The U-937 monocyte-like cell internalizes insulin receptors readily. Incubation with monensin at low doses (10(-6) to 10(-7) M) for 2 h did not affect subsequent surface insulin binding. However, the drug markedly enhanced insulin-induced down-regulation. Monensin had little effect on ligand internalization in this cell line as demonstrated by quantitative morphometric analysis. The IM-9 lymphocyte, a slow internalizer, was less sensitive to monensin exposure. Prolonged exposure (12 h) to this compound of either cell line resulted in apparent inhibition of insertion into the surface membrane of both newly synthesized and recycled receptors. When solubilization was used to quantitate total cell receptors, there was essentially no difference in intact cell binding (i.e. surface receptors) and total cell binding in IM-9 cells when insulin-induced down regulation alone was compared to insulin and monensin. By contrast for the U-937 cells there was only a small further decrease in binding when monensin was added to insulin in the solubilized cells compared to the marked augmentation of down-regulation when monensin was added to insulin in intact cells. These data demonstrate that cells with a rapid internalization rate have an associated active recycling process. By contrast cells with a slow internalization rate have a similarly slow recycling rate. This is consistent with relatively equal rates of receptor biosynthesis and plasma membrane insertion in both cell types.     

Analysis of Effectiveness of Human U-500 Insulin in Patients Unresponsive to Conventional Insulin Therapy

ObjectiveTo present a retrospective analysis of the effects of human U-500 insulin in 20 patients with type 2 diabetes and insulin resistance.MethodsMedical records of 20 patients with type 2 diabetes who had received U-500 insulin for at least 6 months were reviewed to determine glycemic control before and after this therapy. Human regular U-500 insulin therapy was initiated at a unit dosage equivalent to the previous standard insulin dosage. Comparisons of hemoglobin A1c levels, insulin doses, and body mass index at baseline and 6 months after changing to U-500 insulin therapy were analyzed.ResultsIn the 10 female and 10 male study subjects, the mean (± SD) hemoglobin A1c level was 9.59 ± 1.37% initially, and it decreased to 8.53 ± 1.11% at 3 months and to 7.83 ± 1.26% at 6 months after initiation of U-500 insulin therapy. In comparison with baseline, these decreases were statistically significant. The amount of insulin used and the body mass index did not change significantly from baseline to 6 months after initiation of U-500 insulin treatment.ConclusionAnalysis of data suggests that U-500 insulin therapy yields improved glycemic control in insulin-resistant patients who have poor control of blood glucose with use of standard insulin regimens. There is no proof, however, that improved glycemic control by this method decreases diabetes-related complications or improves survival. Further studies must be performed before U-500 insulin can be recommended as a standard therapy for patients with insulin resistance. (Endocr Pract. 2005;11:305-307)     

The Effect of Prestudy Insulin Therapy on Safety and Efficacy of Human Regular U-500 Insulin by Pump or Injection: A Posthoc Analysis

ObjectiveWe conducted a posthoc analysis of the VIVID study (Safety and Efficacy of Human Regular U-500 Insulin Administered by Continuous Subcutaneous Insulin Infusion Versus Multiple Daily Injections in Subjects With Type 2 Diabetes Mellitus: A Randomized, Open-Label, Parallel Clinical Trial), comparing 2 delivery methods of human regular U-500 insulin (U-500R), continuous subcutaneous insulin infusion (CSII) versus multiple daily injection (MDI), in type 2 diabetes requiring high insulin, to determine influence of prestudy insulin on glycemic outcomes.MethodsWe compared A1C, total daily insulin dose (TDD), weight, and hypoglycemia by subgroups of prestudy insulin (prestudy U-500R vs non-U-500R) and treatment (CSII vs MDI).ResultsAt baseline, prestudy U-500R had higher TDD, higher body mass index, lower A1C and fasting plasma glucose, and higher rate of hypoglycemia compared to non-U-500R. Active titration of U-500R reduced A1C in both subgroups, with maximum benefit at 8 weeks. At 26 weeks, CSII provided the greatest reduction in A1C in both subgroups, with a greater reduction in non-U-500R. MDI provided an A1C reduction in both subgroups, with the greater reduction in non-U-500R. At 8 weeks, prestudy U-500R reached its lowest A1C; thereafter, A1C rebounded with MDI and remained stable with CSII. In non-U-500R, A1C continued to decrease to study end. In non-U-500R, hypoglycemia increased during active titration, but then decreased in the posttitration maintenance period. In both subgroups, TDD increased from baseline with MDI but not with CSII. Body weight increased in both subgroups but was greater in prestudy U-500R with CSII compared to MDI.ConclusionRegardless of previous insulin, people on high-dose insulin could lower A1C with U-500R, with additional benefit from CSII. These results may provide guidance for use of U-500R in clinical practice.     

A 24-Week,Prospective, Randomized,Open-Label,Treat-To-Target Pilot Study of Obese Type 2 Diabetes Patients with Severe Insulin Resistance to Assess the Addition of Exenatide on the Efficacy of U-500 Regular Insulin Plus Metformin

ObjectiveTo compare the efficacy of 500 U/mL (U-500) regular insulin + metformin with U-500 regular insulin + metformin + exenatide in improving glycemic control in patients with severely insulin-resistant type 2 diabetes mellitus (T2DM).MethodsThirty patients with T2DM and severe insulin resistance were screened, and 28 were randomized to regular insulin U-500 + metformin or the GLP-1 analog exenatide, U-500, and metformin. Glycated hemoglobin (HbA_1c) levels, body weight, and insulin doses were documented at baseline and at 3 and 6 months. The number and severity hypoglycemic episodes were noted.ResultsThere were 7 males and 7 females in each group (U-500 + metformin and U-500 + metformin + exenatide). Overall, U-500 insulin + metformin, either alone or with the addition of exenatide, resulted in a significant improvement in HbA_1c in both groups, with no significant difference between the 2 groups. There was no meaningful weight change in those utilizing exenatide. Those on U-500 insulin and metformin alone had a tendency toward some weight gain. No severe hypoglycemia occurred during the study period. Symptomatic hypoglycemia was more common in the group on exenatide, but this occurred in only 5 patients, and the clinical significance of this is uncertain. Insulin dosage changes on U-500 regular insulin were variable but tended to be lower in those subjects on exenatide.ConclusionsU-500 regular insulin + metformin is effective for the treatment of T2DM patients with severe insulin resistance. The addition of exenatide may ameliorate potential weight gain but provides no additional improvement in glycemia. (Endocr Pract. 2014;20:1143-1150)     

Glycemic Outcomes of Hospitalized Patients on Ambulatory Humulin-R U-500 Insulin

ObjectiveManaging hospitalized patients on ambulatory U-500 insulin is challenging because of limited guidance on how to safely adjust insulin doses during admission. We sought to evaluate glycemic outcomes in relation to inpatient insulin doses in patients receiving U-500 prior to hospitalization.MethodsRetrospective study of hospitalized patients on ambulatory U-500 seen consecutively from January 2015 to December 2019. Primary outcomes were inpatient hypoglycemia, hyperglycemia, and normoglycemia at different insulin dosages expressed as weight-based (unit/kg/d) inpatient total daily dose (TDD) and ratio of inpatient to outpatient TDD.ResultsWe identified 66 admissions of 46 unique patients. The median (interquartile range) body mass index was 41.0 kg/m² (35.1, 46.8), home TDD 212 units (120, 300), and home insulin dose 1.6 units/kg/d (1.1, 2.2). The median (interquartile range) inpatient insulin dose was 0.7 unit/kg/d (0.3, 1.0) and the ratio of inpatient to outpatient TDD was 0.4 (0.2, 0.8). Hyperglycemia persisted throughout the hospitalization. For the outcomes of hyperglycemia and normoglycemia, we found no association between increased levels of insulin dosages. For the outcome of hypoglycemia, significantly higher odds were observed when non-fasting patients received an inpatient TDD that was either > 40% of their home TDD or > 0.6 unit/kg/d of insulin.ConclusionPatients on ambulatory U-500 have significant hyperglycemia during admission. Inpatient insulin doses of 40% of home TDD or ≤ 0.6 unit/kg were not associated with increased hypoglycemia risk. Further prospective studies are needed to determine effective doses in these high-risk patients.     

Non-plasma bonding of PDMS for inexpensive fabrication of microfluidic devices

In this video, we demonstrate how to use the neuron microfluidic device without plasma bonding. In some cases it may be desirable to reversibly bond devices to the Corning No. 1 cover glass. This could be due, perhaps, to a plasma cleaner not being available. In other instances, it may be desirable to remove the device from the glass after the culturing of neurons for certain types of microscopy or for immunostaining, though it is not necessary to remove the device for immunostaining since the neurons can be stained in the device. Some researchers, however, still prefer to remove the device. In this case, reversible bonding of the device to the cover glass makes that possible. There are some disadvantages to non-plasma bonding of the devices in that not as tight of a seal is formed. In some cases axons may grow under the grooves rather than through them. Also, because the glass and PDMS are hydrophobic, liquids do not readily enter the device making it necessary at times to force media and other reagents into the device. Liquids will enter the device via capillary action, but it takes significantly longer as compared to devices that have been plasma bonded. The plasma cleaner creates temporary hydrophilic charges on the glass and device that facilitate the flow of liquids through the device after bonding within seconds. For non-plasma bound devices, liquid flow through the devices takes several minutes. It is also important to note that the devices to be used with non-plasma bonding need to be sterilized first, whereas plasma treated devices do not need to be sterilized prior to use because the plasma cleaner will sterilize them.     

A simple diluter for flow-through toxicity studies

An original, simple, and inexpensive diluter for flow-through toxicity studies is described. The diluter is run only by water gravity and has no moving parts. The main units of the device are made of very cheap disposable components and may be mounted without qualified technical assistance. It has neither metal nor any other kind of chemically active parts contacting experimental media. The diluter may be easily modified to study toxicants independently or in mixtures, in fact, as many toxicants as are necessary. When properly calibrated, it does not require further adjustments even during long-term experiments. The main original feature of the diluter described here is its dosing system, which is based on a vacuum air lock. This key system is very simple, reliable, includes neither mechanical nor electrical devices, and ensures adequate accuracy in dosing.