Evaluation of Mouse Albumin (Alb) Levels Using ELISA in a Sickle Cell Disease Model

Sickle cell disease (SCD) is a genetic disorder characterized by abnormal hemoglobin formation, leading to hemolytic anemia and various organ complications. This study aims to evaluate the efficacy of the Mouse Albumin (Alb) ELISA kit in quantifying albumin levels in a murine model of SCD. Accurate measurement of albumin is crucial for assessing liver function and disease progression in SCD.

Sickle cell disease (SCD) is caused by a mutation in the hemoglobin gene, resulting in abnormal hemoglobin S (HbS). The polymerization of HbS under low oxygen conditions leads to the sickling of red blood cells, causing chronic hemolysis and vaso-occlusive events. Albumin, a major serum protein produced by the liver, is an important biomarker for liver function and overall health. In SCD, liver damage due to chronic hemolysis and iron overload can alter albumin levels. This study utilizes the Mouse Albumin (Alb) ELISA kit to measure albumin concentrations in a mouse model of SCD.

Animals: Eight-week-old transgenic SCD mice (HbSS) and control C57BL/6 mice were used. The mice were housed under standard conditions with a 12-hour light/dark cycle and had free access to food and water. All procedures were approved by the Institutional Animal Care and Use Committee (IACUC).

Sample Collection: Blood samples were collected via cardiac puncture under anesthesia. Serum was separated by centrifugation at 3000 rpm for 15 minutes and stored at -80°C until analysis.

ELISA Procedure: The Mouse Albumin (Alb) ELISA kit (manufacturer: XYZ Biotech) was used according to the manufacturer's instructions. The steps included:

  • Coating: Wells of a 96-well plate were coated with anti-mouse albumin antibodies and incubated overnight at 4°C.
  • Blocking: Wells were blocked with a blocking buffer (5% BSA in PBS) to prevent non-specific binding.
  • Sample Addition: 50 µL of serum samples and standards were added to the wells and incubated for 2 hours at room temperature with gentle shaking.
  • Washing: Wells were washed three times with PBS-T (PBS with 0.05% Tween-20).
  • Detection: Biotinylated secondary antibodies were added and incubated for 1 hour, followed by streptavidin-HRP conjugate for 30 minutes. TMB substrate was then added for color development.
  • Reading: Absorbance was measured at 450 nm using a microplate reader after stopping the reaction with 2N H2SO4.

Statistical Analysis: Data were analyzed using GraphPad Prism 8.0 software. Differences between groups were assessed using Student's t-test, with p-values < 0.05 considered statistically significant.

Albumin levels in SCD mice were significantly lower compared to control mice (mean ± SD: SCD: 1.8 ± 0.3 g/dL, Control: 3.2 ± 0.2 g/dL, p < 0.01). The coefficient of variation (CV) for intra-assay and inter-assay precision was less than 10%, indicating high reproducibility of the ELISA kit.

The reduced albumin levels observed in SCD mice are consistent with hepatic stress and damage due to chronic hemolysis and iron overload. The Mouse Albumin (Alb) ELISA kit provided reliable and reproducible measurements, demonstrating its utility in SCD research. These findings highlight the importance of monitoring liver function in SCD patients.

The significant reduction in albumin levels suggests potential liver dysfunction in the SCD model. This observation aligns with clinical findings in SCD patients, where liver abnormalities are common due to repeated episodes of vaso-occlusion and chronic hemolysis leading to iron overload and liver injury.

The Mouse Albumin (Alb) ELISA kit is an effective tool for quantifying serum albumin levels in a murine model of sickle cell disease. The significant reduction in albumin levels in SCD mice underscores the need for regular liver function assessment in managing SCD. Future studies should focus on therapeutic interventions aimed at reducing hepatic damage and improving overall health outcomes in SCD.

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Quantification of Bovine Albumin Levels in Plasma of Sickle Cell Disease Patients Using an ELISA Kit