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Total Iron Binding Capacity (TIBC) Colorimetric Assay Kit– MSE Supplies LLC

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Total Iron Binding Capacity (TIBC) Colorimetric Assay Kit

SKU: E-BC-K071-M-96

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Total Iron Binding Capacity (TIBC) Colorimetric Assay Kit

SKU # E-BC-K071-M
Detection Instrument Microplate reader (510-530 nm, optimum wavelength: 520 nm)
Detection Method Colorimetric method


Product Details

Properties

Synonyms TIBC
Sample Type Serum
Sensitivity 0.14 mg/L
Detection Range 0.31-50 mg/L
Detection Method Colorimetric method
Assay type Quantitative
Assay time 20 min
Precision Average inter-assay CV: 2.300% | Average intra-assay CV: 1.500
Other instruments required Micropipettor, Water bath, Incubator, Vortex mixer, Centrifuge
Storage 2-8℃
Valid period 12 months


Dilution of Sample

It is recommended to take 2~3 samples with expected large difference to do pre-experiment before formal experiment and dilute the sample according to the result of the pre-experiment and the detection range (0.31-50 mg/L).

The recommended dilution factor for different samples is as follows (for reference only):

Sample type Dilution factor
Human serum 1
Rat serum 1
Porcine serum 1
Rabbit serum 1
Chicken serum 1
Machin serum 1

 

Note: The diluent is double distilled water.

 

Detection Principle

The excess iron is added to the serum to bind all the ferritin in the serum, and the excess iron is adsorbed by adding the iron adsorbent. The iron bind with the ferritin is separated from the protein by the action of acid solution and reductant. Fe3+ in serum is reduced to Fe2+, Fe2+ binds with bipyridine to form pink complex. In a certain range, the amount of TIBC is positively correlated with the depth of color. The iron content measured is, minus serum iron value, which is called unsaturated iron binding force. Total iron binding capacity minus serum iron value is unsaturated iron binding capacity (UIBC).

 

Kit Components & Storage

Item Component Size 1(48 T) Size 2(96 T) Storage
Reagent 1 100 mg/L Iron Standard Stock Solution 2 mL ×1 vial 2 mL ×1 vial 2-8℃, 12 month
Reagent 2 Chromogenic Agent A Powder × 1 vial Powder × 2 vials 2-8℃, 12 month shading light
Reagent 3 Chromogenic Agent B Powder × 1 vial Powder × 2 vials 2-8℃, 12 month shading light
Reagent 4 Chromogenic Agent C 15 mL × 1 vial 15 mL × 2 vials 2-8℃, 12 month
Reagent 5 Iron Absorbent Powder × 31 vials Powder × 79 vials 2-8℃, 12 month
Microplate 96 wells No requirement
Plate Sealer 2 pieces

 

Note: The reagents must be stored strictly according to the preservation conditions in the above table. The reagents in different kits cannot be mixed with each other. For a small volume of reagents, please centrifuge before use, so as not to obtain sufficient amount of reagents.


Technical Data:

Parameter:

Intra-assay Precision

Three human serum samples were assayed in replicates of 20 to determine precision within an assay (CV = Coefficient of Variation).

Parameters Sample 1 Sample 2 Sample 3
Mean (mg/L) 3.80 24.60 43.80
%CV 1.7 1.6 1.2


Inter-assay Precision

Three human serum samples were assayed 20 times in duplicate by three operators to determine precision between assays.

Parameters Sample 1 Sample 2 Sample 3
Mean (mg/L) 3.80 24.60 43.80
%CV 2.5 2.1 2.3

 

Recovery

Take three samples of high concentration, middle concentration and low concentration to test the samples of each concentration for 6 times parallelly to get the average recovery rate of 100%.


Sample 1 Sample 2 Sample 3
Expected Conc. (mg/L) 8.5 22.6 32
Observed Conc. (mg/L) 8.6 22.4 32.0
Recovery rate (%) 101 99 100


Sensitivity

The analytical sensitivity of the assay is 0.14 mg/L. This was determined by adding two standard deviations to the mean O.D. obtained when the zero standard was assayed 20 times, and calculating the corresponding concentration.

 

Standard Curve

As the OD value of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique or temperature effects), so the standard curve and data are provided as below for reference only:

Concentration (μmol/L) 0 5 10 20 25 30 40 50
Average OD 0.037 0.108 0.178 0.318 0.384 0.454 0.590 0.736
Absoluted OD 0 0.071 0.141 0.281 0.347 0.417 0.553 0.699