QUANTI-Blue™

QUANTI-Blue™ Solution is a colorimetric enzyme assay developed to determine any alkaline phosphatase (AP) activity in a biological sample, such as cell culture supernatant. It has been optimized for use in 96-well plates (standard procedure) and in 1536-well plates (high-throughput screening procedure). 

QUANTI-Blue™ Solution can be used to detect and quantify AP or SEAP activity. Secreted embryonic alkaline phosphatase (SEAP), a truncated form of placental AP, is a widely used reporter gene. SEAP is secreted into cell culture supernatant and therefore offers many advantages over intracellular reporters. InvivoGen provides a wide collection of reporter cell lines with the SEAP reporter gene.

Features and advantages of QUANTI-Blue™ Solution:

• A convenient, easy and rapid means to detect and quantify SEAP and AP activity
• Short hands-on-time: < 10 min
• Easy to prepare: just mix QB Reagent and QB Buffer with water
• Highly sensitive
• Wide dynamic range
• High saturation threshold
• Allows for high-throughput screening (HTS)

In the presence of AP or SEAP, the color of QUANTI-Blue™ changes from pink to purple/blue. The purple/blue color intensity reflects the activity of AP. The levels of AP can be determined qualitatively with the naked eye or quantitatively using a spectrophotometer at 620-655 nm.

Figures

Specifications

Physicochemical characterization: pH, appearance

Functional characterization: cellular assays using alkaline phosphatase of SEAP-expressing reporter cells

Contents

QUANTI-Blue™ Solution is available in three pack sizes:

• rep-qbs : 5 x 1 ml of QB reagent + 5 x 1 ml QB buffer, sufficient to prepare QUANTI‑Blue™ Solution for 25 x 96-well plates or 20 x 1536-well plates
• rep-qbs2 : 10 x 1 ml of QB reagent + 10 x 1 ml QB buffer, sufficient to prepare QUANTI‑Blue™ Solution for 50 x 96-well plates or 40 x 1536-well plates
• rep-qbs3 : 1 x 20 ml of QB reagent + 1 x 20 ml QB buffer, sufficient to prepare QUANTI‑Blue™ Solution for 100 x 96-well plates or 80 x 1536-well plates

Store QB reagent and QB buffer at -20°C.

Reconstituted QUANTI-Blue™ Solution is stable for 2 weeks at 2-8 °C and for 2 months at -20°C. Keep reconstituted QUANTI-Blue™ away from light.

Details

QUANTI-Blue™ Standard procedure

1. Prepare QUANTI-Blue™ Solution as per the instructions on the data sheet. Dispense 180 μl of QUANTI-Blue™ Solution per well of 96-well plate.

2. Add 20 μl of culture supernatant from SEAP-expressing cells to each well.

3. After a 15 min to 6 hour-incubation at 37°C, measure optical density (OD) at 620-655 nm using a microplate reader