LDH-Blue™ Plus
-
Cat.code:
rep-ldhv2-1
- Documents
Note: LDH-Blue™ Plus is a new version of our LDH-Blue™ kit, providing enhanced performance and reagent stability. The cat code has been changed accordingly (rep-ldhv2-1).
ABOUT
Sensitive colorimetric assay for LDH-based cell death detection
InvivoGen introduces LDH-Blue™ Plus, a new and more stable version of its innovative colorimetric assay for measuring lactate dehydrogenase (LDH) activity in cell culture supernatants from damaged or lysed cells. This kit features a substrate that generates a blue formazan product, shifting the medium color from pale red to blue/purple. In contrast to conventional LDH assays that produce red formazan, this blue signal is clearly distinguishable from phenol red in culture media, enabling easy naked-eye detection.
Optimized for simplicity, sensitivity, and accuracy, LDH-Blue™ Plus is ideally suited for cytotoxicity assays, immune cell-mediated killing assays, inflammasome studies, and drug toxicity screening.
Why choose LDH-Blue™ Plus?
The blue formazan readout minimizes interference from phenol red, resulting in a low baseline, high signal clarity, and a wider assay window for precise detection of cytotoxicity.
- Clear visual detection: distinct signal from phenol red
- Low baseline signal: minimized background from spontaneous LDH release
- High linearity: reliable quantification across multiple cell types
- Accurate quantification: comparable performance to fluorescence-based Live/Dead staining
- Broad dynamic range: detection of both low and high levels of cytotoxicity
- Extended reagent stability: storage for up to 1 month at 4 °C and 6 months at -20°C
Assay principle
LDH is a stable cytoplasmic enzyme released into the culture medium upon loss of plasma membrane integrity. LDH-Blue™ Plus detects extracellular LDH via a coupled enzymatic reaction that converts a tetrazolium salt into a blue formazan product, directly proportional to LDH activity.
This color change can be quantified using a spectrophotometer at 650 nm.
Key features
- Add, mix, read: no complex handling steps
- < 30 min assay time: rapid results
- High sensitivity: detection of subtle cytotoxic effects
- Phenol red media compatible: no signal interference
- Cost-efficient: suitable for routine screening
Read our review on the principles and limitations of LDH-based assays
All products are for research use only, and not for human or veterinary use.
| Typical LDH assays | LDH-Blue™ Plus | ||
|---|---|---|---|
| ✖ | Difficult color readout in phenol red medium | ✔ | Clear blue/purple color distinct from phenol red |
| ✖ | Background signal from medium | ✔ | Low background signal from medium |
| ✖ | Higher baseline signal from spontaneous LDH release | ✔ | Low baseline signal |
| ✖ | Reduced dynamic range | ✔ | Larger assay window |
| ✖ | Overestimate cell death | ✔ | Matches flow cytometry Live/Dead™ assay |
| ✖ | Variable reagent stability | ✔ | Stable at 4°C for up to 1 month |
| ✖ | Dry ice shipping | ✔ | RT shipping (economical & environment friendly) |
LDH-Blue™ Plus advantages
LDH-Blue™ Plus assay at a glance
SPECIFICATIONS
Specifications
50 µl (if using P96 well plates)
Each lot is functionally tested and validated.
CONTENTS
Contents
-
Product:LDH-Blue™ Plus
-
Cat code:rep-ldhv2-1
-
Quantity:200 tests
- 1 vial of Reagent Mix (lyophilized)
- 1 bottle (10 ml) Assay Buffer
- 1 bottle (2.5 ml) Lysis Buffer
- 1 bottle (10 ml) Stop Solution
Each vial of Reagent Mix is sufficient for monitoring LDH activity in 2 x 96-well plates (~200 tests).
Shipping & Storage
- Shipping method: Room temperature
- Store the Reagent Mix at -20°C
- Store the Assay Buffer, Lysis Buffer, and Stop Solution at 4°C
- Avoid repeated freeze-thaw cycles
- Protect from light
Storage:
Caution:
Details
Assay principle
The LDH activity is typically determined by utilizing a coupled enzymatic reaction where LDH oxidizes lactate to pyruvate, converting NAD+ to NADH. Subsequent reduction of tetrazolium salts by NADH and a phenazine derivative electron mediator leads to the formation of colored formazan. The amount of formazan is directly proportional to cell viability.
InvivoGen's LDH-Blue™ kit contains all the necessary compounds for assessing LDH activity, including the substrate, which, upon reduction, produces blue formazan. It can be easily detected with the naked eye and quantified by measuring the absorbance at 650 nm with a spectrophotometer.
Illustration created with Biorender.com.
DOCUMENTS
Documents
Technical Data Sheet
Validation Data Sheet
Safety Data Sheet
Certificate of analysis
Need a CoA ?
You may also need
Frequently Asked Questions
LDH-Blue™
To monitor cytotoxicity using InvivoGen's LDH-Blue™ Plus, you will need:
- test and control supernatants
- pipettes and pipette tips, including multi-channel pipette
- reagent reservoir
- flat-bottom transparent 96-well plates
- a spectrophotometer microplate reader capable of reading 650nm absorbance.
The kit components are shipped at room temperature. Store the Reagent Mix at -20 °C and protect it from light. Store the Assay Buffer, Lysis Buffer, and Stop Solution at 4 °C. When stored as directed, the kit is stable for up to 1 year from the date of receipt. Once resuspended, the Reagent Mix must be kept protected from light and is stable for up to 1 month at 4°C, and for up to 6 months at -20 °C. Avoid repeated (>3) freeze-thaw cycles.
LDH-Blue™ Plus detects lytic cell death associated with plasma membrane disruption, including necroptosis, pyroptosis, ferroptosis, late apoptosis, and immune cell-mediated cytotoxicity. It is not specific to apoptotic pathways without membrane rupture.
LDH-Blue™ Plus primarily detects cell death associated with membrane integrity loss, such as necrosis or late-stage apoptosis. For early apoptosis detection, complementary assays like Annexin V staining are recommended.
Absolutely. LDH-Blue™ Plus is ideal for T cell, NK cell, or antibody-mediated cytotoxicity assays. Ensure proper controls for spontaneous and maximum release.
Yes, LDH-Blue™ Plus is sensitive enough to detect LDH release from samples with low cell numbers, including 3D microtissues and primary cells. Its high sensitivity ensures accurate cytotoxicity assessment in various experimental setups.
Absolutely. LDH-Blue™ Plus is designed for use with standard 96-well and 384-well plate formats, making it suitable for high-throughput screening of cytotoxicity in drug discovery and other large-scale studies. You must change the volumes of the supernatant and reagent mix solution (and stop solution) according to the plate format. For a 96-well plate, please see the technical datasheet.
Yes, LDH-Blue™ Plus is optimized for use with serum-containing or serum-free media. Background levels may vary slightly with serum type.
Certain substances, such as high concentrations of reducing agents or components that affect enzyme activity, may interfere with the assay. It's advisable to include appropriate controls to account for potential interferences.
Depending on your assay, you may want to store the supernatants for later reading. We recommend storing the supernatant at 4°C for up to 48 hours and at -80°C for a longer period. Do not store at -20°C.
There are different possible explanations for low absorbance values:
- The LDH reaction time is too short. If so, do not use the optional Stop Solution and perform kinetic readings of LDH activity.
- The cell culture density is too low. If so, repeat the experiment to optimize cell number.
- The test compound may interfere with the LDH oxydo-reductive reaction (see FAQ above).
- Bubbles are present in the wells when performing the reading. Avoid bubble formation when pipetting and mixing.
One possible reason for the high variation in replicate values could be pipetting errors or insufficient mixing of the Reagent Mix Solution with supernatant samples. We recommend you repeat the experiment using another set of pipettes and/or mix the plate more thoroughly (avoiding bubble formation).
Since LDH-Blue™ Plus is non-radioactive and uses absorbance-based detection, it can be multiplexed with fluorescence or luminescence-based assays from the same sample (with appropriate timing). For example, you can use LDH-Blue™ Plus with InvivoGen's Lucia luciferase-based cellular assays: the HMGB1 release reporter cells THP1-HMGB1-Lucia™, the ferroptosis reporter cells HT1080-HMGB1-Lucia™, and the oxidative stress reporter cells HEK-Lucia-Star™ ARE.
Powered by Bioz