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Rat Serine / threonine-protein kinase STK11 / LKB1 ELISA Kit

SKU:
RTFI00359
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
D4AE59
Sensitivity:
0.094ng/ml
Range:
0.156-10ng/ml
ELISA Type:
Sandwich
Synonyms:
STK11, PJS, EC 2.7.11.1, LKB1 serine, threonine kinase 11, Peutz-Jeghers syndrome, PJS, PJS polarization-related protein LKB1, Renal carcinoma antigen NY-REN-19, serine, threonine kinase 11, serine, threonine-protein kinase 11, Serine, threonine-prot
Reactivity:
Rat
Research Area:
Cell Death
€649
Frequently bought together:

Description

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Product Name:Rat STK11 (Serine/threonine-protein kinase STK11) ELISA Kit
Product Code:RTFI00359
Size:96 Assays
Target:Rat STK11
Alias:STK11, PJS, EC 2.7.11.1, LKB1 serine, threonine kinase 11, Peutz-Jeghers syndrome, PJS, PJS polarization-related protein LKB1, Renal carcinoma antigen NY-REN-19, serine, threonine kinase 11, serine, threonine-protein kinase 11, Serine, threonine-protein kinase LKB1
Reactivity:Rat
Detection Method:Sandwich ELISA, Double Antibody
Sensitivity:0.094ng/ml
Range:0.156-10ng/ml
Storage:4°C for 6 months
Note:For Research Use Only
Recovery:Matrices listed below were spiked with certain level of Rat STK11 and the recovery rates were calculated by comparing the measured value to the expected amount of Rat STK11 in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)85-10395
EDTA plasma(n=5)87-10496
UFH plasma(n=5)86-10595
Linearity:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Rat STK11 and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)89-104%89-100%87-103%
EDTA plasma(n=5)83-101%83-95%82-99%
UFH plasma(n=5)80-98%82-95%80-97%
Intra-Assay:CV <8%
Inter-Assay:CV <10%
Uniprot:D4AE59
UniProt Protein Function:Tumor suppressor serine/threonine-protein kinase that controls the activity of AMP-activated protein kinase (AMPK) family members, thereby playing a role in various processes such as cell metabolism, cell polarity, apoptosis and DNA damage response. Acts by phosphorylating the T-loop of AMPK family proteins, thus promoting their activity: phosphorylates PRKAA1, PRKAA2, BRSK1, BRSK2, MARK1, MARK2, MARK3, MARK4, NUAK1, NUAK2, SIK1, SIK2, SIK3 and SNRK but not MELK. Also phosphorylates non-AMPK family proteins such as STRADA, PTEN and possibly p53/TP53. Acts as a key upstream regulator of AMPK by mediating phosphorylation and activation of AMPK catalytic subunits PRKAA1 and PRKAA2 and thereby regulates processes including: inhibition of signaling pathways that promote cell growth and proliferation when energy levels are low, glucose homeostasis in liver, activation of autophagy when cells undergo nutrient deprivation, and B-cell differentiation in the germinal center in response to DNA damage. Also acts as a regulator of cellular polarity by remodeling the actin cytoskeleton. Required for cortical neuron polarization by mediating phosphorylation and activation of BRSK1 and BRSK2, leading to axon initiation and specification. Involved in DNA damage response: interacts with p53/TP53 and recruited to the CDKN1A/WAF1 promoter to participate in transcription activation. Able to phosphorylate p53/TP53; the relevance of such result in vivo is however unclear and phosphorylation may be indirect and mediated by downstream STK11/LKB1 kinase NUAK1. Also acts as a mediator of p53/TP53-dependent apoptosis via interaction with p53/TP53: translocates to the mitochondrion during apoptosis and regulates p53/TP53-dependent apoptosis pathways. In vein endothelial cells, inhibits PI3K/Akt signaling activity and thus induces apoptosis in response to the oxidant peroxynitrite. Regulates UV radiation-induced DNA damage response mediated by CDKN1A. In association with NUAK1, phosphorylates CDKN1A in response to UV radiation and contributes to its degradation which is necessary for optimal DNA repair.
UniProt Code:D4AE59
NCBI GenInfo Identifier:347662427
NCBI Gene ID:314621
NCBI Accession:D4AE59.1
UniProt Secondary Accession:D4AE59,D4A179,
UniProt Related Accession:D4AE59
Molecular Weight:46,758 Da
NCBI Full Name:Serine/threonine-protein kinase STK11
NCBI Synonym Full Names:serine/threonine kinase 11
NCBI Official Symbol:Stk11  
NCBI Official Synonym Symbols:Lkb1  
NCBI Protein Information:serine/threonine-protein kinase STK11
UniProt Protein Name:Serine/threonine-protein kinase STK11
UniProt Synonym Protein Names:Liver kinase B1 homolog; LKB1
Protein Family:Serine/threonine-protein kinase
UniProt Gene Name:Stk11  
StepProcedure
1. Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells!
2.Aliquot 0.1ml standard solutions into the standard wells.
3. Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well.
4. Add 0.1 ml of properly diluted sample ( Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells.
5.Seal the plate with a cover and incubate at 37°C for 90 min.
6. Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2.
7. Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall.
8.Seal the plate with a cover and incubate at 37°C for 60 min.
9. Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash.
10. Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min.
11. Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min.
12. Add 90 µL of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color.
13. Add 50 µL of Stop solution into each well and mix thoroughly. The color changes into yellow immediately.
14. Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.

Sample Type Protocol
Serum:

If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

If serum separator tubes are not being used, allow samples to clotovernight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Removeserum and assay promptly or aliquot and store the samples at-80°C. Avoid multiple freeze-thaw cycles.

Plasma: Collect plasma using EDTA or heparin as an anti-coagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.Note: Over haemolysed samples are not suitable for use with this kit.
Urine & Cerebrospinal Fluid: Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.
Cell Culture Supernatant: Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.
Cell Lysates: Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20°C.
Tissue Homogenates: The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenizein 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or-80°C.
Tissue Lysates: Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Breast Milk: Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.

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