You are here : Home / Products / Biomarkers / Biomarkers / Apolipoprotein assays

Apolipoprotein assays: ApoA1, ApoB, ApoC3, ApoE

Apolipoproteins HTRF kit

Homogeneous, cell-based quantification of human apolipopoproteins

The HTRF® apolipoprotein assays provide all necessary reagents for rapid and  sensitive quantification of human apolipoprotein A1, B, C3 or E in a homogeneous format. The assays may be used with cell culture supernatants, whole cells, lipoproteins and plasma. The assays are easily adaptable for 96- to 384-w formats and enables higher throughput compared to ELISA .

Apolipoproteins are constituents of triglyceride-rich lipoprotein including VLDL (or bad cholesterol) and HDL (or good cholesterol). They play a central role in regulating triglyceride metabolism and are key markers of cardiovascular diseases and  type 2 diabetes.

Features

  • Results in less than 3 hours
  • Ready to use kits: detecting reagents, standard and buffers provided
  • Simple "mix and read" protocol
  • Low volume sample size needed: <10µL
  • Automation & high throughput compatible
  • Extremely robustness with cell culture media

 

Applications

  • Quantification of apolipoprotein in human cell supernatants, whole cells, in lipoproteins and plasma
  • Screening modulators of apolipoprotein production
  • Monitoring nuclar receptor activity
  • Cardiovascular, Diabetes, CNS

 

The HTRF Apolipoprotein assays are sandwich immunoassays involving two monoclonal antibodies, one labelled with Eu-Cryptate (Donor) and the second with d2  (Acceptor). The intensity of the signal is proportional to the concentration of apolipoprotein present in the sample.

All four apolipoprotein assays are based on the same assay protocol and can be run with cell supernatants or whole cells, lipoproteins and human plasma. The antibodies labelled with HTRF fluorophores may be pre-mixed and added in a single dispensing step, to further streamline the protocol. The assay can be run in 96- to 384-well plates by simply resizing each addition volume proportionally. The figure on the right shows the Apo A1 assay protocol in a 384-well small volume white plate (20 µL final). When using the cell-based procedure, the detecting reagents are directly added to the assay plate following the stimultion step.

 

 

Cell-based protocol in 384w

 

  Human Apo A1 Human Apo B Human Apo CIII Human Apo E
Incubation 2h at RT 3h at RT 2h at RT (or ON) 3h at RT (or ON)
Assay range 0 to 4000 ng/mL 0 to 4000 ng/mL 8 to 125 ng/mL 0.5 to 32 ng/mL
Detection limit 0.5 ng/mL 1.8 ng/mL 2.0 ng/mL 0.9 ng/mL
Extended range Up to 8,000 ng/ml Up to 8,000 ng/ml NA Up to 100 ng/ml
Cross reactivity

Low cross reactivity with: ApoB-100, ApoA2,

ApoC1, ApoC2, ApoC3, ApoE

Low cross reactivity with: ApoA1, ApoA2,

ApoC1, ApoC2, ApoC3, ApoE

Crossreactivity with ApoB-48: 100%

NA NA

(*) Based on 384-well low volume plate (20µL). Extending incubation period improves sensitivity

 

Cross-reactivity


Cross-reactivity (%) ApoA1 ApoB
ApoA1 100 0.004
ApoB-100 0.044 100
ApoA2 0.037 <0.001
ApoC1 0.014 <0.001
ApoC2 0.004 0.001
ApoC3 0.005 0.006
ApoE 0.004 0.004
ApoB-48 ND 100

 

 

 

Cell-based apolipoprotein assays to monitor Retinoid X receptor (RXR) agonist:

Human hepatoma cells (HepG2) in 80-90% confluence were grown in DMEM (Gibco; 11885, low glucose) supplemented with 2% FBS. 5 μL of HepG2 cells (2,000 & 5,000 per well) and 5 μL of the RXR ligand 9-cis retinoic acid (9-cis-RA) in DMEM,  2% FBS at various concentrations were dispensed into a 384-well low volume black culture plate. After a 48 hour incubation at 37°C in CO2 atmosphere, 10 μL of the two pre-mixed HTRF® detection reagents (from either the ApoA1 or the ApoB assay kit) were added. Readout was performed on RUBYstar after 3 hours of incubation at room temperature (an extract from the poster by K Adachi).

Ordering Info

DescriptionCat. noProduct insertMSDS
Human Apolipoprotein C3 kit - 1,000 tests63ADK001PEB
-
Human Apolipoprotein C3 kit - 20,000 tests63ADK001PEC
-
Human Apolipoprotein E kit - 1,000 tests63ADK004PEB
-
Human Apolipoprotein E kit - 20,000 tests63ADK004PEC
-
Human Apolipoprotein A1 kit - 1,000 tests64APAPEB
-
-
Human Apolipoprotein A1 kit - 20,000 tests64APAPEC
-
-
Human Apolipoprotein B kit - 1,000 tests64APBPEB
-
-
Human Apolipoprotein B kit - 20,000 tests64APBPEC
-
-




© 2013 Cisbio.com - All Rights Reserved