Overview

  • Product name

    LDL Uptake Assay Kit (Cell-Based)
  • Detection method

    Fluorescent
  • Sample type

    Adherent cells
  • Assay type

    Cell-based
  • Assay time

    6h 30m
  • Species reactivity

    Reacts with: Mammals, Other species
  • Product overview

    LDL Uptake Assay Kit (Cell-Based) (ab133127) provides a convenient tool for studying LDL uptake and regulation at the cellular level. This kit employs Human LDL conjugated to DyLight™ 550 as a fluorescent probe for detection of LDL uptake into cultured cells. An LDL receptor-specific polyclonal antibody and a DyLight™ 550-conjugated secondary antibody are included for identifying the distribution of LDL receptors.


    LDL uptake assay protocol summary:
    - remove culture medium from experimentally treated cells
    - add LDL-Dylight 550 solution and incubate for 3-4 hrs
    - replace solution with culture medium
    - analyze LDL uptake with fluorescent microscope
    - wash cells and  fix for 10 min
    - wash 3 times for 5 min
    - add assay blocking solution and incubate for 30 min
    - add anti-LDL receptor antibody and incubate for 1 hr
    - wash 3 times for 5 min
    - add Dylight 488 secondary antibody and incubate for 1 hr
    - wash 3 times for 5 min
    - analyze staining with fluorescence microscope

  • Notes

    LDL is the major carrier of cholesterol in the blood, accounting for more than 60% of total plasma cholesterol. LDL is taken up by hepatic and extra-hepatic tissues through receptor mediated endocytosis triggered by apoB-100-LDL receptor interaction. The internalized LDL particle is transported to lysosomes where it is degraded to free cholesterol and amino acids.

  • Platform

    Fluorescence microscope

Properties

Images

  • LDL uptake in pre-adipocytes and adipocytes.

    Panel A: Light image of undifferentiated pre-adipocytes.
    Panel B: Undifferentiated pre-adipocytes do not show any LDL uptake.
    Panel C: Undifferentiated pre-adipocytes show little LDL receptor expression.
    Panel D: Light image of differentiated adipocytes.
    Panel E: Corresponding cells to those in Panel D showing uptake of LDL (red).
    Panel F: Corresponding cells to those in Panel D showing expression of LDL receptor (green).
  • LDL uptake in HepG2 cells.
    HepG2 cells were cultured at a density of 3 x 104 cells/well in a 96 well plate for two days then treated with 32.5µM EGCG overnight. LDL-Dylight™ 549 (10&microg/ml) was added and the cells were incubated for an additional four hours. Cells were fixed and stained for LDL receptor using a Rabbit anti-LDL receptor primary antibody and DylightTM 488-conjugated secondary antibody.
    Panel A: LDL-Dylight™ 549 taken into cells appears in red.
    Panel B: LDL receptors (in green) show a distribution pattern that matches the cells in the Panel A containing LDL-Dylight™ 549.

Protocols

References

This product has been referenced in:

  • Peng WC  et al. Inflammatory Cytokine TNFa Promotes the Long-Term Expansion of Primary Hepatocytes in 3D Culture. Cell 175:1607-1619.e15 (2018). Read more (PubMed: 30500539) »
  • Qian T  et al. Directed differentiation of human pluripotent stem cells to blood-brain barrier endothelial cells. Sci Adv 3:e1701679 (2017). Human . Read more (PubMed: 29134197) »
See all 5 Publications for this product

Customer reviews and Q&As

Answer



I can confirm you that the concentration of LDL-DYLight 549 stock solution supplied in the LDL uptake assay kit (ab133127) is 1 mg/mL.

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