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Protocol for Albuwell M kit: Murine Microalbuminuria ELISA By Exocell Inc Resource Report Resource Website |
Kathi Burke, Peter Reifsnyder | DOI:10.17504/protocols.io.34mgqu6 | Diabetic Complications Consortium | Albuwell M is an indirect competitive ELISA designed to monitor kidney function in the mouse by measurement of urinary albumin. To complete the assay, sample and rabbit anti-murine albumin antibody are added to albumin coated wells. The antibody interacts and binds with the albumin immobilized to the stationary phase or with albumin in the fluid phase, hence the notion of competitive binding. A subsequent reaction with anti-rabbit -HRP conjugate labels the probe with enzyme. After washing, only the antibody-conjugate bound to the stationary phase remains in the well, and this is detected using a chromogenic reaction. Color intensity is inversely proportional to the logarithm of albumin in the fluid phase. The assay may be completed in less than 2.5 hoursDiabetic Complication: | University of Arizona, Frank Brosius Lab, The Jackson Laboratory, Ed Leiter Lab | https://www.diacomp.org/shared/document.aspx?id=22&docType=Protocol | 1 | 2019 | Kathi Burke, Peter Reifsnyder 2019. Protocol for Albuwell M kit: Murine Microalbuminuria ELISA By Exocell Inc. protocols.io https://dx.doi.org/10.17504/protocols.io.34mgqu6 | 2021-04-15 09:15:37 | ||
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Biochemical Measures of Neuropathy - GSSG (Non-Enzymatic) Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.3qpgmvn | Diabetic Complications Consortium | Summary:Oxidative stress is highly correlated with the metabolic changes caused by hyperglycemia. Increased levels of glucose overload mitochondria and result in the production of reactive oxygen species (ROS). In addition, the flow of excess glucose through cellular pathways decreases the cell’s normal ability to detoxify ROS. As a result, the neurons and axons of the peripheral nervous system contain increased levels of ROS and decreased antioxidant capacity. The following assays are used to measure these changes in rodent models of diabetic neuropathy. Diabetic Complication: | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=54&docType=Protocol | 1 | 2019 | Eva Feldman 2019. Biochemical Measures of Neuropathy - GSSG (Non-Enzymatic). protocols.io https://dx.doi.org/10.17504/protocols.io.3qpgmvn | 2021-04-15 09:15:56 | ||
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Neuropathy Phentoyping Protocols - Dosages of Analgesics for Rats and Mice Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.3jrgkm6 | Diabetic Complications Consortium | Summary:Phenotyping of Rodents for the Presence of Diabetic Neuropathy In man, the development of diabetic neuropathy is dependent on both the degree of glycemic control and the duration of diabetes. Diabetic neuropathy is a progressive disorder, with signs and symptoms that parallel the loss of nerve fibers over time. Consequently, assessments of neuropathy in mice are not performed at one time point, but are characterized at multiple time points during a 6 month period of diabetes. The degree of diabetes is evaluated in 2 ways: tail blood glucose measured following a 6 hour fast and glycated hemoglobin levels. The initial degree of neuropathy is screened using the methods discussed below. Detailed measures of neuropathy are employed when the initial screening instruments indicate a profound or unique phenotypic difference. This document contains protocols used by the DiaComp staff to examine and measure diabetic neuropathy at the whole animal, tissue and cellular levels.Diabetic Complication: | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=54&docType=Protocol | 1 | 2019 | Eva Feldman 2019. Neuropathy Phentoyping Protocols - Dosages of Analgesics for Rats and Mice. protocols.io https://dx.doi.org/10.17504/protocols.io.3jrgkm6 | 2021-04-15 09:15:59 | ||
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Frequently sampled Insulin glucose tolerance test Resource Report Resource Website |
Timothy Nichols, David Clemmons | DOI:10.17504/protocols.io.8e8hthw | Diabetic Complications Consortium | This assay is used by the DiaComp to measure glucose tolerance and insulin sensitivity in pigs.Diabetic Complications:Reference:1. Bergman RN, Finegood DT, Ader M: Assessment of insulin sensitivity in vivo. Endocr Rev 1985, 6:45-86. | University of North Carolina at Chapel Hill, University of North Carolina at Chapel Hill | https://www.diacomp.org/shared/document.aspx?id=50&docType=Protocol | 2 | 2019 | Timothy Nichols, David Clemmons 2019. Frequently sampled Insulin glucose tolerance test. protocols.io https://dx.doi.org/10.17504/protocols.io.8e8hthw | 2021-04-15 09:16:09 | ||
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Neuropathy Phentoyping Protocols - Hematoxylin and Eosin Staining Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.3jsgkne | Diabetic Complications Consortium | Summary:Phenotyping of Rodents for the Presence of Diabetic Neuropathy In man, the development of diabetic neuropathy is dependent on both the degree of glycemic control and the duration of diabetes. Diabetic neuropathy is a progressive disorder, with signs and symptoms that parallel the loss of nerve fibers over time. Consequently, assessments of neuropathy in mice are not performed at one time point, but are characterized at multiple time points during a 6 month period of diabetes. The degree of diabetes is evaluated in 2 ways: tail blood glucose measured following a 6 hour fast and glycated hemoglobin levels. The initial degree of neuropathy is screened using the methods discussed below. Detailed measures of neuropathy are employed when the initial screening instruments indicate a profound or unique phenotypic difference. This document contains protocols used by the DiaComp staff to examine and measure diabetic neuropathy at the whole animal, tissue and cellular levels.Diabetic Complication:Reference: AFIP Laboratory Methods in Histotechnology, 1994, p. 16, 53-55. | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=54&docType=Protocol | 1 | 2019 | Eva Feldman 2019. Neuropathy Phentoyping Protocols - Hematoxylin and Eosin Staining. protocols.io https://dx.doi.org/10.17504/protocols.io.3jsgkne | 2021-04-15 09:16:09 | ||
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Tail Flick Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.7thhnj6 | Diabetic Complications Consortium | The Tail Flick assay is a pain receptive assay in which a mouse is placed within a restraining tube with its tail protruding. The tail is placed on a level surface, a radiant heat is applied to the tail and the latency of the mouse to remove its tail from the heat is recorded. This latency is used as a measure to indicate neurological pathology.Diabetic Complication: | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=34&docType=Protocol | 2 | 2019 | Eva Feldman 2019. Tail Flick. protocols.io https://dx.doi.org/10.17504/protocols.io.7thhnj6 | 2021-04-15 09:16:12 | ||
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Neuropathy Phentoyping Protocols - Immunohistochemistry for Tissue Sections Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.7rfhm3n | Diabetic Complications Consortium | Summary:Phenotyping of Rodents for the Presence of Diabetic Neuropathy In man, the development of diabetic neuropathy is dependent on both the degree of glycemic control and the duration of diabetes. Diabetic neuropathy is a progressive disorder, with signs and symptoms that parallel the loss of nerve fibers over time. Consequently, assessments of neuropathy in mice are not performed at one time point, but are characterized at multiple time points during a 6 month period of diabetes. The degree of diabetes is evaluated in 2 ways: tail blood glucose measured following a 6 hour fast and glycated hemoglobin levels. The initial degree of neuropathy is screened using the methods discussed below. Detailed measures of neuropathy are employed when the initial screening instruments indicate a profound or unique phenotypic difference. This document contains protocols used by the DiaComp staff to examine and measure diabetic neuropathy at the whole animal, tissue and cellular levels.Diabetic Complication: | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=54&docType=Protocol | 2 | 2019 | Eva Feldman 2019. Neuropathy Phentoyping Protocols - Immunohistochemistry for Tissue Sections. protocols.io https://dx.doi.org/10.17504/protocols.io.7rfhm3n | 2021-04-15 09:16:10 | ||
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Induce Stroke via Hypoxia-Ischemia Resource Report Resource Website |
Ian Simpson | DOI:10.17504/protocols.io.8e5htg6 | Diabetic Complications Consortium | This document describes the procedure for inducing a stroke in diabetic RCS10 mice, their non-diabetic controls and NON/ShiLtJ control mice. Our initial objective was to use those RCS10 mice that did not become diabetic as the controls for the experiment to compare the effect of diabetes on stroke outcome and that data is illustrated in Figure 2. However, with further breading the numbers of non- diabetic RCS 10 mice at twenty weeks of age were so few that it became necessary to use the NON/ShiLtJ mice which are 85% genetically identical to the RCS10 mice and stroke outcome is illustrated in Figure 3.Diabetic Complications: | Pennsylvania State University | https://diacomp.org/shared/document.aspx?id=251&docType=Protocol | 2 | 2019 | Ian Simpson 2019. Induce Stroke via Hypoxia-Ischemia. protocols.io https://dx.doi.org/10.17504/protocols.io.8e5htg6 | 2021-04-15 09:16:13 | ||
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Quantitative PCR analysis to assess gene expression changes in hyperglycemic larval zebrafish Resource Report Resource Website |
Sandra Rieger | DOI:10.17504/protocols.io.3bagiie | Diabetic Complications Consortium | To assess hyperglycemia in larval zebrafish, we analyzed the expression of genes implicated in glucose metabolism, such as insulin, insulin receptor, glucagon, and phosphoenolpyruvate carboxylase (pepck), using quantitative PCR (qPCR). We analyzed gene expression following treatment of Tg(ins:NTRmCherry) transgenic larvae either with 0.5 % DMSO (controls) or 10 mM metronidazole (see Protocol 2) for β-cell ablation at 3 and 8 days post fertilization (dpf). Diabetic Complication: | University of Miami | https://www.diacomp.org/shared/document.aspx?id=222&docType=Protocol | 1 | 2019 | Sandra Rieger 2019. Quantitative PCR analysis to assess gene expression changes in hyperglycemic larval zebrafish. protocols.io https://dx.doi.org/10.17504/protocols.io.3bagiie | 2021-04-15 09:16:11 | ||
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Biochemical Measures of Neuropathy - Western Blot Stripping Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.3s3gngn | Diabetic Complications Consortium | Summary:Oxidative stress is highly correlated with the metabolic changes caused by hyperglycemia. Increased levels of glucose overload mitochondria and result in the production of reactive oxygen species (ROS). In addition, the flow of excess glucose through cellular pathways decreases the cell’s normal ability to detoxify ROS. As a result, the neurons and axons of the peripheral nervous system contain increased levels of ROS and decreased antioxidant capacity. The following assays are used to measure these changes in rodent models of diabetic neuropathy. Diabetic Complication: | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=54&docType=Protocol | 1 | 2019 | Eva Feldman 2019. Biochemical Measures of Neuropathy - Western Blot Stripping. protocols.io https://dx.doi.org/10.17504/protocols.io.3s3gngn | 2021-04-15 09:16:28 | ||
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MOVAT Pentachrome Staining Resource Report Resource Website |
Charles Alpers | DOI:10.17504/protocols.io.346gqze | Diabetic Complications Consortium | The following table describes the staining color for various structures.Results: Nuclei and Elastic Fibers Black Ground Substance and Mucin Blue Muscle Red Collagen and Reticular Fibers Yellow Fibrinioid and Fibrin Intense RedDiabetic Complications: | University of Washington | https://www.diacomp.org/shared/document.aspx?id=75&docType=Protocol | 1 | 2019 | Charles Alpers 2019. MOVAT Pentachrome Staining. protocols.io https://dx.doi.org/10.17504/protocols.io.346gqze | 2021-04-15 09:16:17 | ||
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Biochemical Measures of Neuropathy - Glutathione Reductase Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.3qsgmwe | Diabetic Complications Consortium | Summary:Oxidative stress is highly correlated with the metabolic changes caused by hyperglycemia. Increased levels of glucose overload mitochondria and result in the production of reactive oxygen species (ROS). In addition, the flow of excess glucose through cellular pathways decreases the cell’s normal ability to detoxify ROS. As a result, the neurons and axons of the peripheral nervous system contain increased levels of ROS and decreased antioxidant capacity. The following assays are used to measure these changes in rodent models of diabetic neuropathy. Diabetic Complication: | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=54&docType=Protocol | 1 | 2019 | Eva Feldman 2019. Biochemical Measures of Neuropathy - Glutathione Reductase. protocols.io https://dx.doi.org/10.17504/protocols.io.3qsgmwe | 2021-04-15 09:16:25 | ||
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TRAP Assay Resource Report Resource Website |
Eva Feldman | DOI:10.17504/protocols.io.3sqgndw | Diabetic Complications Consortium | Summary:The use of total radical-trapping antioxidant parameter (TRAP) has recently been proposed to explore the antioxidant property of a plasma sample. This assay is a measure of oxidative stress in the animals. This protocol describes the procedure used by the DiaComp to measure TRAP.Diabetic Complications: | University of Michigan - Ann Arbor | https://www.diacomp.org/shared/document.aspx?id=32&docType=Protocol | 1 | 2019 | Eva Feldman 2019. TRAP Assay. protocols.io https://dx.doi.org/10.17504/protocols.io.3sqgndw | 2021-04-15 09:16:50 | ||
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Vascular perfusion of mice Resource Report Resource Website |
Peter Mundel | DOI:10.17504/protocols.io.8ezhtf6 | Diabetic Complications Consortium | This protocol describes the procedure for perfusion of mice prior to organ or tissue harvesting .Diabetic Complications: | Massachusetts General Hospital, Boston, MA | https://diacomp.org/shared/document.aspx?id=24&docType=Protocol | 2 | 2019 | Peter Mundel 2019. Vascular perfusion of mice. protocols.io https://dx.doi.org/10.17504/protocols.io.8ezhtf6 | 2021-04-15 09:16:59 | ||
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Breeding Scheme and Selection of Animals for DiaComp Experiments Resource Report Resource Website |
Nobuyo Maeda, Oliver Smithies, Nobuyuki Takahashi | DOI:10.17504/protocols.io.8fphtmn | Diabetic Complications Consortium | SummaryIt is extremely important to measure quantitative parameters (e.g. plasma glucose, urinary albumin, etc.) in a way that minimizes the effects of any adventitious variables, such as seasonal changes, dietary differences, any differences in the degrees of backcrossing, or unplanned differences in genotypes. This means that, whenever possible, mice of the critical genotypes should be obtained from genotypically identical matings carried out at or close to the same time. Archival data are not appropriate. The breeding scheme to generate the experimental animals can vary in complexity, depending on the fertility of the two genders, whether the Testgene is to be studied in homozygotes or in heterozygotes, and whether the diabetes is induced genetically or by STZ. However, it is important to remember that, when looking for the effects of the Testgene genotype on diabetic complications, the aim is to compare the Testgene mutant/mutant (or Testgene mutant/wt) diabetic animals with their diabetic litter mates that are wildtype at the Testgene locus (Testgene wt/wt). ReferenceWang, C-H, Li, F, Hiller, S, Kim, H-S, Maeda, N, Smithies, O, and Takahashi, N. A modest decrease in endothelial NOS in mice comparable to that associated with human NOS3 variants exacerbates diabetic nephropathy. Proc. Natl. Acad. Sci. USA 108(5): 2070-2075 (2011) PMID: 21245338 PMCID: PMC3033253.Diabetic Complications: | University of North Carolina at Chapel Hill, University of North Carolina at Chapel Hill, Tohoku University | https://www.diacomp.org/shared/document.aspx?id=79&docType=Protocol | 2 | 2019 | Nobuyo Maeda, Oliver Smithies, Nobuyuki Takahashi 2019. Breeding Scheme and Selection of Animals for DiaComp Experiments. protocols.io https://dx.doi.org/10.17504/protocols.io.8fphtmn | 2021-04-15 09:16:59 | ||
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Mouse dissection and preparation of heart (aortic sinus) and brachiocephalic artery for lesion quantitation by sectioning Resource Report Resource Website |
Jan Breslow, Daniel Teupser | DOI:10.17504/protocols.io.3a5gig6 | Diabetic Complications Consortium | Diabetic complication: | Rockefeller University, Rockefeller University | https://www.diacomp.org/shared/document.aspx?id=13&docType=Protocol | 1 | 2019 | Jan Breslow, Daniel Teupser 2019. Mouse dissection and preparation of heart (aortic sinus) and brachiocephalic artery for lesion quantitation by sectioning. protocols.io https://dx.doi.org/10.17504/protocols.io.3a5gig6 | 2021-04-15 09:16:41 | ||
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En face preparation and quantification of aortic surface area covered by atherosclerotic lesions Resource Report Resource Website |
Jan Breslow, Daniel Teupser | DOI:10.17504/protocols.io.8fehtje | Diabetic Complications Consortium | This protocol describes how the DiaComp members prepare the En Face staining for atherosclerotic lesions quantification.Diabetic Complication: | The Rockefeller University, The Rockefeller University | https://www.diacomp.org/shared/document.aspx?id=12&docType=Protocol | 2 | 2019 | Jan Breslow, Daniel Teupser 2019. En face preparation and quantification of aortic surface area covered by atherosclerotic lesions. protocols.io https://dx.doi.org/10.17504/protocols.io.8fehtje | 2021-04-15 09:16:49 | ||
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Quantitative Magnetization Transfer MR Imaging (qMT-MRI) of Diabetic Mouse Kidney Resource Report Resource Website |
Feng Wang, Takamune Takahashi | DOI:10.17504/protocols.io.8a7hshn | Diabetic Complications Consortium | This protocol describes the procedures and data acquisition of quantitative magnetization MR imaging of diabetic mouse kidney.Reference: 1. Zhao, H.J., et al., Endothelial nitric oxide synthase deficiency produces accelerated nephropathy in diabetic mice. J Am Soc Nephrol, 2006. 17(10): p. 2664-2669. 2. Ramani, A., et al., Precise estimate of fundamental in-vivo MT parameters in human brain in clinically feasible times. Magnetic resonance imaging, 2002. 20(10): p. 721-31. 3. Cercignani, M. and G.J. Barker, A comparison between equations describing in vivo MT: the effects of noise and sequence parameters. Journal of Magnetic Resonance, 2008. 191(2): p. 171-83. 4. Pluim, J.P.W., J.B.A. Maintz, and M.A. Viergever, Mutual-information-based registration of medical images: A survey. Ieee Transactions on Medical Imaging, 2003. 22(8): p. 986- 1004. 5. Wang, F., et al., Repeatability and sensitivity of high resolution blood volume mapping in mouse kidney disease. J Magn Reson Imaging, 2014. 39(4): p. 866-71. 6. Smith, S.A., et al., Measurement of T1 and T2 in the cervical spinal cord at 3 tesla. Magnetic resonance in medicine : official journal of the Society of Magnetic Resonance in Medicine / Society of Magnetic Resonance in Medicine, 2008. 60(1): p. 213-9. 7. Henkelman, R.M., et al., Quantitative Interpretation of Magnetization-Transfer. Magnetic Resonance in Medicine, 1993. 29(6): p. 759-766. Diabetic Complication: | Vanderbilt University, Vanderbilt University | https://www.diacomp.org/shared/document.aspx?id=303&docType=Protocol | 2 | 2019 | Feng Wang, Takamune Takahashi 2019. Quantitative Magnetization Transfer MR Imaging (qMT-MRI) of Diabetic Mouse Kidney. protocols.io https://dx.doi.org/10.17504/protocols.io.8a7hshn | 2021-04-15 09:17:03 | ||
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Morphometry: Mouse Resource Report Resource Website |
Ed Leiter | DOI:10.17504/protocols.io.7rhhm36 | Diabetic Complications Consortium | This protocol describes a number of the measurements that are made on the mouse anatomy.Edited by: DiaComp MembershipDiabetic Complications: | THE JACKSON LABORATORY | https://www.diacomp.org/shared/document.aspx?id=47&docType=Protocol | 2 | 2019 | Ed Leiter 2019. Morphometry: Mouse. protocols.io https://dx.doi.org/10.17504/protocols.io.7rhhm36 | 2021-04-15 09:16:55 | ||
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High Dose STZ Induction Protocol Resource Report Resource Website |
Frank Brosius | DOI:10.17504/protocols.io.8iyhufw | Diabetic Complications Consortium | This protocol is used by DiaComp members to induce diabetes in a number of the animal models developed by the consortium. STZ is toxic to the insulin- producing beta cells of the pancreas and used to induce a diabetes similar to a type I diabetic (Reference). Some reports suggest cellular toxicity outside of the pancreas. STZ also exhibits broad spectrum antibacterial properties and alters the gut microbiota. Please ensure that appropriate controls are included in all studies and complementary models considered (e.g. the Ins2Akita mouse).Diabetic Complications: | University of Arizona | https://www.diacomp.org/shared/document.aspx?id=74&docType=Protocol | 2 | 2019 | Frank Brosius 2019. High Dose STZ Induction Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.8iyhufw | 2021-04-15 09:17:08 |
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