Human Galectin-3 ELISA kit

Human Galectin-3 ELISA kit

SKU: 31690

 

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Assay range : 0.625-40 ng/mL

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  • INTRODUCTION

    Galectin-3 is a member of the b-galactoside-binding lectins family which has conserved carbohydrate recognition domain (CDR). It is a 35 kDa protein located in cytoplasm, nucleus and extracellular spaces. It is also also a member of the beta-galactoside-binding protein family that plays an important role in cell-cell adhesion, cell matrix interactions, macrophage activation, angiogenesis, apoptosis and insulin resistance. Elevated serum galectin-3 levels were observed in Behcet’s disease, thyroid, Alzheimer’s disease, cardiovascular disease such as left atrial appendage (LAA) stroke and in several types of cancers especially when it is metastatic. Moreover, circulating levels of galectin-3 were higher in obese patients and it is an indicator of insulin resistance.

  • PRINCIPLE OF THE ASSAY

    This assay is a quantitative sandwich ELISA. The immunoplate is pre-coated with a polyclonal antibody specific for human galectin-3. Standards and samples are pipetted into the wells and any human galectin-3 present is bound by the immobilized antibody. After washing away any unbound substances, a horseradish peroxidase (HRP)-linked polyclonal antibody specific for human galectin-3 is added to the wells. After a final wash step, an HRP substrate solution is added and colour develops in proportion to the amount of human galectin-3 bound initially. The assay is stopped and the optical density of the wells determined using a microplate reader. Since the increases in absorbance are directly proportional to the amount of captured human galectin-3, the unknown sample concentration can be interpolated from a reference curve included in each assay.

  • ASSAY PERFORMANCE

    A. Typical representation of standard curve

    The following standard curve is provided for demonstration only. A standard curve should be generated for each set of sample assay. 

    Human galectin-3 (ng/ml)

    Absorbance

    (450 nm)

    Blanked Absorbance

    0

    0.082

    0

    0.625

    0.111

    0.029

    1.25

    0.15

    0.068

    2.5

    0.213

    0.131

    5

    0.404

    0.357

    10

    0.782

    0.70

    20

    1.542

    1.46

    40

    2.663

    2.581

     

    B. Sensitivity:

    The lowest level of human galectin-3 that can be measured by this assay is 0.145 ng/ml.

          

    C. Specificity:

    Cross reactivity of recombinant proteins:

    Analyte

    Cross Reactivity

    Human FABP4

    No

    Human LCN2

    No

    Human Adiponectin

    No

    Human FGF-21

    No

     

    D. Precision:

    Intra-assay Precision (Precision within an assay)

    Two samples of known concentration were tested 8 times on one plate.

    Sample

    Mean (ng/ml)

    SD (ng/ml)

    CV (%)

    1

    29.65

    1.33

    4.5

    2

    2.26

    0.14

    6.2

    Inter-assay Precision (Precision between assays)

    Two samples of known concentration were tested in 8 separate assays.

    Sample

    Mean (ng/ml)

    SD (ng/ml)

    CV (%)

    1

    29.5

    1.64

    5.5

    2

    2.31

    0.14

    6.1

     

    E. Spike

    Serum samples were assayed by adding 90 µl of sample and 10 µl of spike stock solution calculated to yield the intended 0, 2.5, 20 or 40 ng/ml spike concentration

    Spike level

    Expected

    (ng/ml)

    Observed

    (ng/ml)

    Recovery (%)

    Low spike (2.5 ng/ml)

    2.23

    2.02

    90.3

    Medium spike (20 ng/ml)

    23.1

    17.3

    74.9

    High spike

    (40 ng/ml)

    43.5

    32.9

    75.6

     

    F. Linearity:

    To assess the linearity of the assay, samples containing and/or spiked with high concentrations of human galectin-3 were serially diluted with the 1×Assay buffer to produce samples with values within the dynamic range of the assay.

    Sample1:

    Dilution

    Measured (ng/ml)

    Expected (ng/ml)

    Recovery (%)

    1/2

    35.7

    40

    89.2

    1/4

    35.5

    40

    88.6

    1/8

    36.2

    40

    90.5

     

    G. Validation:

      Serum galectin-3 levels were measured in three different groups, women with normal glucose tolerance during pregnancy (NGT), gestational diabetes mellitus (GDM) and healthy women (age 20-35,Healthy) respectively.

     

     

     

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