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ULK1 ELISA Kit (Unc-51 Like Kinase 1)

Full Name: ULK1 ELISA Kit (Unc-51 Like Kinase 1)
Reactivity: Human
Sample Type: Plasma, Biological Fluids, Serum, Tissue Homogenates
Sensitivity: 9.375 pg/ml

INTRODUCTION

ULK1 is a vital protein during the process of autophagy in many mammalian cells. It is found to be in the activated state during periods of nutrient deprivation, the initiation of autophagy is triggered by many upstream signal pathways. Also, many downstream targets have been identified that can be phosphorylated, for example beclin-1 at the Ser 14 position. This leads to the activation of pro-autophagy class III PI(3)K and VPS34 complex, this results in the promotion of autophagy maturation and induction.

ULK1 is composed of a 112 kDa protein, that contains an interacting domain (C-terminal), a serine-proline rich region and a kinase domain (N-terminal). The serine-proline rich region has been found to be a site for AMPK and mTORC1 phosphorylation. AMPK is a positive regular whereas mTORC1 is a negative regular. There is also evidence to indicate that Unc-51 like kinase 1 may have a positively charged activation loop, this region is found to regulate the activity of the kinase and is vital in helping to recognise various substrates.

The Unc-51 like kinase family has a conserved domain of unknown function, DUF246. This region is usually found in other protein kinases and has no well defined function. However, this region has been shown to be important for the activity of Unc-51 like kinase 1 by “de novo” protein synthesis that was observed upon substrate stimulation. The presence of a conserved domain also helps to reduce variability in the activity of the protein kinase as less variability can be introduced into the response through expression or mutation within this region. The generally accepted mechanism of these kinases is the phosphorylation of a conserved threonine residue to yield the active form. The substrate specificity and activity of Unc-51 like kinase involves a conserved serine at position 502 with only minor differences in various models. The Arg525-His526 loop is essential for kinase activation by tyrosine phosphorylation in response to heparin binding on its receptor.

INTENDED USE

Human ULK1 ELISA kit can detect levels of Unc-51 like kinase 1 (ATG1, ULK1, UNC51) using human serum, tissue homogenates, plasma and other biological samples.

CONTENT

All reagents supplied need to be stored at 2 °C – 8 °C, unopened reagents will retain reactivity until expiration date. Do not use reagents beyond this date.

  • One 96-Well Plate: Pre-coated with anti-ULK1 antibody.
  • Standards: Lyophilized recombinant.
  • Sample/Standard Dilution Buffer.
  • Biotinylated-labelled Antibody.
  • Antibody Dilution Buffer.
  • HRP-Streptavidin Conjugate (SABC).
  • SABC Dilution Buffer.
  • TMB Substrate.
  • Wash Buffer (25x).
  • Plate Sealer.
  • Product Instructions.

TYPICAL RESULTS

For this ULK1 ELISA kit it is recommended that a standard curve is generated for each assay carried out.

Standard Curve: 0, 15.625, 31.25, 62.5, 125, 250, 500, 1000 pg/ml.
Reactivity: Human
Sensitivity: 9.375 pg/ml
Range: 15.625 – 1000 pg/ml
Principle: Sandwich
Application: Research Use Only.

ASSAY CHARACTERISTICS

– Specificity: Highly specific for ULK1, no cross reactivity or interference between ULK1 and analogues was detected.
– Recovery: Serum (87 – 104%), EDTA Plasma (89 – 102%), Heparin Plasma (88 – 103%).
– Linearity: Serum (83 – 101%), EDTA Plasma (83 – 96%), Heparin Plasma (85 – 98%).
– Precison Intra-Assay: CV < 8%.
– Precison Inter-Assay: CV < 10%.
– Stability: Less than 10%.

REFERENCES

  1. MPK and mTOR regulate autophagy through direct phosphorylation of Ulk1. Nat Cell Biol. (2011) 13 (2): 132-41. Kim J., et al.
  2. The mammalian_ULK1 complex and autophagy initiation. Essays Biochem. (2017) 61 (6): 585-596. Zachari M. and Ganley I.G.
  3. DAPK3 inhibits gastric cancer progression via activation of ULK1-dependent autophagy. Cell Death Differ. (2021) 28 (3): 952-967. Li G.M., et al.
  4. Human ULK1-Variation and Susceptibility to Mycobacterium tuberculosis Infection. J Infect Dis. (2016) 214 (8): 1260-7. Horne D.J., et al.
  5. Function and regulation of ULK1: From physiology to pathology. Gene. (2022) 840: 146772. Rong Z., et al.

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