Rat GS (Gelsolin) CLIA Kit (RTES00235)

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RTES00235
€649

Description

ELISA Kit Technical ManualMSDS

Rat GS (Gelsolin) 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 GS . 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 GS 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 GS, 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 GS. The concentration of Rat GS 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 range3.13-200 ng/mL
Sensitivity1.88 ng/mL
Sample volume100uL
Sample typeSerum, plasma and other biological fluids
RepeatabilityCV < 15%

Specificity

This kit recognizes Rat GS in samples. No significant cross-reactivity or interference between Rat GS 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
20041810
46336
4407344035
10016796
19626
1821118173
508669
7743
82068168
253662
4208
39353897
12.52064
1900
19821944
6.251094
1010
10521014
3.13567
629
598560
038
38
38--

Precision

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

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

  Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)9.7223.0876.599.2622.8672.33
Standard deviation1.261.706.411.102.716.94
C V (%)12.967.378.3711.8811.859.59

Recovery

The recovery of Rat GS 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)91-10697
EDTA plasma (n=5)89-10195
Cell culture media (n=5)93-105100

Linearity

Samples were spiked with high concentrations of Rat GS 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 (%)98-11295-10792-106
Average (%)10410198
1:4Range (%)100-11685-9798-113
Average (%)10791104
1:8Range (%)102-118100-11796-109
Average (%)109108102
1:16Range (%)99-11199-11390-105
Average (%)10510797

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 GS (Gelsolin) CLIA Kit (RTES00235) 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 GS (Gelsolin) CLIA Kit (RTES00235) Protein Information

UniProt Protein Function:Gelsolin: a calcium-regulated, actin-modulating protein that binds to the plus (or barbed) ends of actin monomers or filaments, preventing monomer exchange (end-blocking or capping). It can promote the assembly of monomers into filaments (nucleation) as well as sever filaments already formed. Its expression is significantly decreased in many tumors and transformed cell lines. Gelsolin and other proteins including E-cadherin, BRCA, p21WAF, HoxA5, and retinoic acid receptor II are regulated by epigenetic changes in breast cancers. Gelsolin helps mediate the ability of HDAC inhibitors to protect neurons from oxygen/glucose deprivation. Two human isoforms are produced by alternative initiation.
UniProt Protein Details:

Protein type:Motility/polarity/chemotaxis; Secreted; Actin-binding; Secreted, signal peptide; Nuclear receptor co-regulator

Cellular Component: actin cap; actin cytoskeleton; cortical actin cytoskeleton; cytoplasm; cytosol; extracellular region; extracellular space; focal adhesion; lamellipodium; myelin sheath; nucleus; perinuclear region of cytoplasm; plasma membrane; podosome; protein complex; ruffle; sarcoplasm

Molecular Function:actin binding; calcium ion binding; myosin II binding; protein domain specific binding

Biological Process: actin filament capping; actin filament polymerization; actin filament severing; actin nucleation; actin polymerization and/or depolymerization; aging; apoptosis; barbed-end actin filament capping; extracellular matrix disassembly; negative regulation of virion penetration into host cell; oligodendrocyte development; phagocytosis, engulfment; phosphoinositide-mediated signaling; positive regulation of actin nucleation; protein destabilization; regulation of cell adhesion; response to ethanol; response to folic acid; sequestering of actin monomers; striated muscle atrophy; tissue regeneration; vesicle-mediated transport; wound healing

NCBI Summary:plays a role in phosphoinositide mediated disassembly of actin filaments in Sertoli cell adhesion complexes; may regulate sperm release and turnover of the blood-testis barrier [RGD, Feb 2006]
UniProt Code:Q68FP1
NCBI GenInfo Identifier:51854227
NCBI Gene ID:296654
NCBI Accession:NP_001004080.1
UniProt Related Accession:Q68FP1
Molecular Weight:80,929 Da
NCBI Full Name:gelsolin
NCBI Synonym Full Names:gelsolin
NCBI Official Symbol:Gsn  
NCBI Protein Information:gelsolin
UniProt Protein Name:Gelsolin
UniProt Synonym Protein Names:Actin-depolymerizing factor; ADF; Brevin
Protein Family:Gelsolin
UniProt Gene Name:Gsn  
UniProt Entry Name:GELS_RAT
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Additional Information

Product type:
ELISA
Reactivity:
Rat
ELISA Type:
CLIA
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