Rat SP-B (Pulmonary Surfatcant-Associated Protein B) CLIA Kit (RTES00049)

SKU:
RTES00049
Product type:
ELISA
Reactivity:
Rat
ELISA Type:
CLIA
€649

Description

ELISA Kit Technical ManualMSDS

Rat SP-B (Pulmonary Surfatcant-Associated Protein B) CLIA Kit

This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Rat SP-B . Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Rat SP-B and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Rat SP-B, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Rat SP-B. The concentration of Rat SP-B in the samples can be calculated by comparing the RLU of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityRat
Detection methodChemiluminescence
Detection range0.31-20 ng/mL
Sensitivity0.19 ng/mL
Sample volume100uL
Sample typeSerum, plasma and other biological fluids
RepeatabilityCV < 15%

Specificity

This kit recognizes Rat SP-B in samples. No significant cross-reactivity or interference between Rat SP-B and analogues was observed.

Typical Data

As the RLU values of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.

Concentration
(ng/mL)
RLU Average Corrected
2031431
36533
3398233950
1011539
13401
1247012438
55670
4830
52505218
2.52364
2684
25242492
1.251385
1381
13831351
0.63870
864
867835
0.31609
637
623591
032
32
32--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Rat SP-B were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Rat SP-B were tested on 3 different plates, 20 replicates in each plate.

  Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)1.072.248.741.102.448.87
Standard deviation0.110.210.640.110.170.58
C V (%)10.289.387.3210.006.976.54

Recovery

The recovery of Rat SP-B spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type Range (%) Average Recovery (%)
Serum (n=5)98-113104
EDTA plasma (n=5)88-10495
Cell culture media (n=5)101-119108

Linearity

Samples were spiked with high concentrations of Rat SP-B and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.

    Serum (n=5) EDTA plasma (n=5) Cell culture media (n=5)
1:2Range (%)84-9793-10890-104
Average (%)9010196
1:4Range (%)91-10285-9793-105
Average (%)969199
1:8Range (%)95-10685-96102-116
Average (%)10091109
1:16Range (%)92-106101-11488-102
Average (%)9810795

Kit Components & Storage

An unopened kit can be stored at 4'C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.

Item Specifications Storage
Micro CLIA Plate(Dismountable)8 wells X12 strips-20'C, 6 months
Reference Standard2 vials
Concentrated Biotinylated Detection Ab (100X)1 vial, 120 uL
Concentrated HRP Conjugate (100X)1 vial, 120 uL-20'C(shading light), 6 months
Reference Standard & Sample Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25X)1 vial, 30 mL
Substrate Reagent A1 vial, 5 mL4'C (shading light)
Substrate Reagent B1 vial, 5 mL4'C (shading light)
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Rat SP-B (Pulmonary Surfatcant-Associated Protein B) CLIA Kit (RTES00049) Assay procedure

    1. Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
    2. Aliquot 100µl of standard solutions into the standard wells.
    3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids.) into test sample wells.
    5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
    6. Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix.Incubate for 1 hour at 37°C.
    7. Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
    8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
    9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
    10. Add 100µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37°C. Protect the plate from light.
    11. Determine the RLU value of each well immediately.

Rat SP-B (Pulmonary Surfatcant-Associated Protein B) CLIA Kit (RTES00049) Protein Information

UniProt Protein Function:Pulmonary surfactant-associated proteins promote alveolar stability by lowering the surface tension at the air-liquid interface in the peripheral air spaces. SP-B increases the collapse pressure of palmitic acid to nearly 70 millinewtons per meter.
NCBI Summary:acts to lower surface tension in lung alveoli [RGD, Feb 2006]
UniProt Code:P22355
NCBI GenInfo Identifier:149036401
NCBI Gene ID:192155
NCBI Accession:EDL91019.1
UniProt Related Accession:P22355
Molecular Weight:41,590 Da
NCBI Full Name:surfactant associated protein B
NCBI Synonym Full Names:surfactant protein B
NCBI Official Symbol:Sftpb  
NCBI Official Synonym Symbols:Sp-b  
NCBI Protein Information:pulmonary surfactant-associated protein B
UniProt Protein Name:Pulmonary surfactant-associated protein B
UniProt Synonym Protein Names:Pulmonary surfactant-associated proteolipid SPL(Phe)
UniProt Gene Name:Sftpb  
UniProt Entry Name:PSPB_RAT
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