Insulin Assay
Insulin, a peptide hormone, is secreted by pancreatic ß-cells and is a key regulator in glucose homeostasis. Insulin deficiency leads to insulin-dependent (type 1) diabetes, whereas resistance to insulin action is common in non-insulin-dependent diabetes (type 2), obesity, and endocrine dysfunctions. The HTRF® Insulin assay is a homogeneous assay for rapid and accurate measurement of insulin in rat, mouse, pig and human samples. This ready-to-use kit is available with two standard ranges. The choice of the protocol will depend on the nature of the samples:
- High Range protocol (10 to 100 ng/mL) for samples from glucose perfused pancreas
- Sensitive Range protocol (0.2 to 10 ng/mL) for pancreatic ß-cell supernatants and serum samples
- Homogeneous assay
- Assay results in under 3 hours
- Suited for HTS
- Excellent reproducibility: CV<4%
- Small sample volume needed (as low as 5-10 µL)
- Two standard ranges: High Range (10-100 ng/mL) and Sensitive Range (0.2-10 ng/mL)
Glucagon (see assays on demand)
- Insulin measurement
- Screening of insulinotropic molecules
- Diabetes assessment
Assay Principle:
The HTRF® Insulin assay can be used to measure rat, pig, mouse or human insulin in a variety of media including cell supernatants and serum. The assay is a sandwich immunoassay that uses two monoclonal antibodies, one labeled with Eu3+-Cryptate and one labeled with XL665 that recognize distinct epitopes. The assay is performed in a single step and only requires one incubation. The two detection reagents can be premixed and pre-plated, and the plates frozen for later use. This feature is possible due to the stability and robustness of HTRF® reagents, and represents a practical advantage for screening.
Product Specifications:
| Detection limit: | 0.20 ng/mL (Sensitive Range protocol) |
| 4 ng/mL (High Range protocol) |
|
| Dynamic range: | 0.3 to 10 ng/mL (Sensitive Range protocol) |
| 3 to 100 ng/mL (High Range protocol) |
|
| S/B: | 18 in 96- or 384-well plates |
| Z: | as high as 0.89 |
| Specificity: | rat, mouse, porcine & human insulin |
Figure 1: Insulin standard curves obtained with the Sensitive Range protocol and performed using the designated final assay volumes and plate types.
Assay Performance:
Rat and mouse models are commonly used in diabetes research. In these models, insulin secretion is monitored on pancreatic cell lines as well as in biological fluids. Cisbio Bioassays's HTRF insulin assay has been validated using a number of experimental systems including screening of insulinotropic molecules on pancreatic β-cells under HTS conditions and monitoring of in vivo insulin and glucose production.
Pancreatic β-cells
INS-1E pancreatic β-cells underwent starvation for 48 hours prior to stimulation with glucose and additional
insulinotropic agents. Insulin concentrations in cell supernatants were determined using the HTRF Insulin assay following the Sensitive Range protocol.
As shown in figure 2, glucose alone stimulated measurable insulin
levels. As expected, the addition of insulinotropic agents stimulated increased insulin production when
compared to glucose alone.
In vivo Studies
In these studies rats were anesthetized and two catheters inserted, one used to obtain blood samples and the
other used for the injection of glucose. Blood samples were taken at the time of injection and at various
intervals thereafter. Samples were then tested for glucose and insulin levels (Figure 3). Induction of
insulin was easily detected and correlated with glucose levels using as little as 50µl blood samples.
Figure 2: 104 INS-1E pancreatic β-cells /100 µL/well were plated in a 96-well tissue-culture treated plate. Cells were stimulated with 1mM glucose in combination with the compounds designated below the graph. 50 µL of cell supernatants were tested using standard HTRF insulin assay conditions.
Figure 3: Male Wistar rats were injected via catheter with glucose (3g/kg).T=0 is the time of glucose injection. Samples were dispensed into a 96 half-well plate (50 µL/well) and insulin levels measured using the standard HTRF insulin assay protocol.
Ordering Info:
Insulin Assay Reagents:
| Description | Quantity* | Cat no. | Product Insert | ||
| Insulin | 1,000 tests 220 tests** |
62INSPEB | |||
| Insulin bulk | 20,000 tests 4,400 tests** |
62INSPEC | |||
| *based on 20 µl assay volume | |||||
** Kit working with two standard ranges : 0.2 to 10 ng/mL allowing 1,000 and 20,000 tests and 10 to 100 ng/mL allowing 220 and 4,400 tests
Companion Products:
| Description | Cat no. |
| Insulin calibrator | 62INSCDA |
| Insulin control | 62INSTDA |
For more information or To Place an Order:
Europe and other countries
+33(0)466 796 705
