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LMO3 Colorimetric Cell-Based ELISA

SKU:
CBCAB00388
Product Type:
ELISA Kit
ELISA Type:
Cell Based
Research Area:
Epigenetics and Nuclear Signaling
Reactivity:
Human
Mouse
Rat
Detection Method:
Colorimetric
€599
Frequently bought together:

Description

system_update_altDatasheetMSDS

LMO3 Colorimetric Cell-Based ELISA

The LMO3 Colorimetric Cell-Based ELISA Kit is a powerful tool for researchers looking to study the function of LMO3 in cellular processes. This kit allows for the accurate and sensitive detection of LMO3 levels in cell lysates and tissue homogenates, providing valuable insights into the role of LMO3 in various biological pathways.LMO3 is a transcriptional regulator known to be involved in developmental processes and cell differentiation. Dysregulation of LMO3 has been implicated in various diseases, including cancer and neurological disorders, making it a promising target for therapeutic intervention.

With its high sensitivity and specificity, the LMO3 Colorimetric Cell-Based ELISA Kit offers researchers a reliable and reproducible method for quantifying LMO3 levels in their samples. Whether studying the impact of LMO3 on cell signaling pathways or exploring its therapeutic potential, this kit is a valuable addition to any research toolkit.

Product Name:LMO3 Colorimetric Cell-Based ELISA
Product Code:CBCAB00388
ELISA Type:Cell-Based
Target:LMO3
Reactivity:Human, Mouse, Rat
Dynamic Range:> 5000 Cells
Detection Method:Colorimetric 450 nmStorage/Stability:4°C/6 Months
Format:96-Well Microplate

The LMO3 Colorimetric Cell-Based ELISA Kit is a convenient, lysate-free, high throughput and sensitive assay kit that can detect LMO3 protein expression profile in cells. The kit can be used for measuring the relative amounts of LMO3 in cultured cells as well as screening for the effects that various treatments, inhibitors (ie siRNA or chemicals), or activators have on LMO3.

Qualitative determination of LMO3 concentration is achieved by an indirect ELISA format. In essence, LMO3 is captured by LMO3-specific primary antibodies while the HRP-conjugated secondary antibodies bind the Fc region of the primary antibody. Through this binding, the HRP enzyme conjugated to the secondary antibody can catalyze a colorimetric reaction upon substrate addition. Due to the qualitative nature of the Cell-Based ELISA, multiple normalization methods are needed:

1. A monoclonal antibody specific for human GAPDH is included to serve as an internal positive control in normalizing the target absorbance values.
2. Following the colorimetric measurement of HRP activity via substrate addition, the Crystal Violet whole-cell staining method may be used to determine cell density. After staining, the results can be analysed by normalizing the absorbance values to cell amounts, by which the plating difference can be adjusted.
Database Information:Gene ID: 55885, UniProt ID: Q8TAP4, OMIM: 180386, Unigene: Hs.504908
Gene Symbol:LMO3
Sub Type:None
UniProt Protein Function:LMO3: 3 isoforms of the human protein are produced by alternative splicing
UniProt Protein Details:

Chromosomal Location of Human Ortholog: 12p12.3

Cellular Component: cytoplasm

Molecular Function:protein binding; zinc ion binding

Biological Process: positive regulation of fat cell differentiation; regulation of transcription, DNA-dependent; transcription, DNA-dependent

NCBI Summary:The protein encoded by this gene belongs to the rhombotin family of cysteine-rich LIM domain oncogenes. This gene is predominantly expressed in the brain. Related family members, LMO1 and LMO2 on chromosome 11, have been reported to be involved in chromosomal translocations in T-cell leukemia. Many alternatively spliced transcript variants have been found for this gene. [provided by RefSeq, Aug 2011]
UniProt Code:Q8TAP4
NCBI GenInfo Identifier:34098603
NCBI Gene ID:55885
NCBI Accession:Q8TAP4.1
UniProt Secondary Accession:Q8TAP4,Q58A66, Q58A67, Q8N974, Q9UDD5, B4DG90, B4DH35
UniProt Related Accession:Q8TAP4
Molecular Weight:18,438 Da
NCBI Full Name:LIM domain only protein 3
NCBI Synonym Full Names:LIM domain only 3
NCBI Official Symbol:LMO3  
NCBI Official Synonym Symbols:RBTN3; RHOM3; RBTNL2; Rhom-3  
NCBI Protein Information:LIM domain only protein 3
UniProt Protein Name:LIM domain only protein 3
UniProt Synonym Protein Names:Neuronal-specific transcription factor DAT1; Rhombotin-3
Protein Family:LIM domain only protein
UniProt Gene Name:LMO3  
UniProt Entry Name:LMO3_HUMAN
ComponentQuantity
96-Well Cell Culture Clear-Bottom Microplate2 plates
10X TBS24 mL
Quenching Buffer24 mL
Blocking Buffer50 mL
15X Wash Buffer50 mL
Primary Antibody Diluent12 mL
100x Anti-Phospho Target Antibody 60 µL
100x Anti-Target Antibody60 µL
Anti-GAPDH Antibody60 µL
HRP-Conjugated Anti-Rabbit IgG Antibody12 mL
HRP-Conjugated Anti-Mouse IgG Antibody12 mL
SDS Solution12 mL
Stop Solution24 mL
Ready-to-Use Substrate12 mL
Crystal Violet Solution12 mL
Adhesive Plate Seals2 seals

The following materials and/or equipment are NOT provided in this kit but are necessary to successfully conduct the experiment:

  • Microplate reader able to measure absorbance at 450 nm and/or 595 nm for Crystal Violet Cell Staining (Optional)
  • Micropipettes with capability of measuring volumes ranging from 1 µL to 1 ml
  • 37% formaldehyde (Sigma Cat# F-8775) or formaldehyde from other sources
  • Squirt bottle, manifold dispenser, multichannel pipette reservoir or automated microplate washer
  • Graph paper or computer software capable of generating or displaying logarithmic functions
  • Absorbent papers or vacuum aspirator
  • Test tubes or microfuge tubes capable of storing ≥1 ml
  • Poly-L-Lysine (Sigma Cat# P4832 for suspension cells)
  • Orbital shaker (optional)
  • Deionized or sterile water

*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.

StepProcedure
1.Seed 200 µL of 20,000 adherent cells in culture medium in each well of a 96-well plate. The plates included in the kit are sterile and treated for cell culture. For suspension cells and loosely attached cells, coat the plates with 100 µL of 10 µg/ml Poly-L-Lysine (not included) to each well of a 96-well plate for 30 minutes at 37°C prior to adding cells.
2.Incubate the cells for overnight at 37°C, 5% CO2.
3.Treat the cells as desired.
4.Remove the cell culture medium and rinse with 200 µL of 1x TBS, twice.
5.Fix the cells by incubating with 100 µL of Fixing Solution for 20 minutes at room temperature. The 4% formaldehyde is used for adherent cells and 8% formaldehyde is used for suspension cells and loosely attached cells.
6.Remove the Fixing Solution and wash the plate 3 times with 200 µL 1x Wash Buffer for five minutes each time with gentle shaking on the orbital shaker. The plate can be stored at 4°C for a week.
7.Add 100 µL of Quenching Buffer and incubate for 20 minutes at room temperature.
8.Wash the plate 3 times with 1x Wash Buffer for 5 minutes each time.
9.Add 200 µL of Blocking Buffer and incubate for 1 hour at room temperature.
10. Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
11.Add 50 µL of 1x primary antibodies (Anti-LMO3 Antibody and/or Anti-GAPDH Antibody) to the corresponding wells, cover with Parafilm and incubate for 16 hours (overnight) at 4°C. If the target expression is known to be high, incubate for 2 hours at room temperature.
12.Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
13.Add 50 µL of 1x secondary antibodies (HRP-Conjugated AntiRabbit IgG Antibody or HRP-Conjugated Anti-Mouse IgG Antibody) to corresponding wells and incubate for 1.5 hours at room temperature.
14.Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
15.Add 50 µL of Ready-to-Use Substrate to each well and incubate for 30 minutes at room temperature in the dark.
16.Add 50 µL of Stop Solution to each well and read OD at 450 nm immediately using the microplate reader.

(Additional Crystal Violet staining may be performed if desired – details of this may be found in the kit technical manual.)

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