Lactate Dehydrogenase B / LDH-B Activity Assay (ab140361)
Key features and details
- Assay type: Enzyme activity
- Detection method: Colorimetric
- Platform: Microplate reader
- Sample type: Adherent cells, Cell culture supernatant, Cell Lysate, Plasma, Purified protein, Serum, Tissue Extracts
Overview
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Product name
Lactate Dehydrogenase B / LDH-B Activity Assay
See all Lactate Dehydrogenase B/LDH-B kits -
Detection method
Colorimetric -
Precision
Intra-assay Sample n Mean SD CV% Overall 3 5% Inter-assay Sample n Mean SD CV% Overall 4 5% -
Sample type
Cell culture supernatant, Serum, Plasma, Adherent cells, Tissue Extracts, Cell Lysate, Purified protein -
Assay type
Enzyme activity -
Range
8 ng/ml - 500 ng/ml -
Recovery
Sample specific recovery Sample type Average % Range Serum 95 86% - 111% Cell culture media 94 91% - 101% -
Species reactivity
Reacts with: Goat, Cow, Human
Does not react with: Mouse, Rat -
Product overview
The microplate assay ab140361 is used to determine Lactate dehydrogenase B / LDH-B activity (LDHB) in a sample utilizing the immunocapture technique. The specific native LDHB enzyme is immune-captured within the wells of the microplate and activity is determined by following the production of NADH catalyzed by the enzyme. The generation of NADH is coupled to the 1:1 reduction of a reporter dye to yield a colored (yellow) reaction product whose concentration can be monitored by measuring the increase in absorbance at 450 nm (Dye molar extinction coefficient - 37000 M-1 cm-1). The immune-capture based activity assay kit contains all the required reagents for a fast and simple measurement of LDHB activity in a whole cell extract, tissue homogenate, or serum.
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Notes
Functional lactate dehydrogenase are homo or hetero tetramers composed of M and H protein subunits encoded by the LDHA and LDHB genes, respectively. The major isozymes of skeletal muscle and liver has four muscle (M) subunits; while the main isozymes for heart muscle contains four (H) subunits. The other variants contain both types of subunits. Elevated levels of LDH indicate tissue breakdown in hemolysis. LDH can be used as a marker of myocardial infarction, peaking at days 3 and lasting through day 10. Other disorders indicated by elevated LDH include cancer, meningitis, encephalitis, acute pancreatitis, and HIV. Defects in LDHB are a cause of hereditary LDHB deficiency; however LDHB deficiency is usually asymptomatic.
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Platform
Microplate reader
Properties
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Storage instructions
Store at +4°C. Please refer to protocols. -
Components 1 x 96 tests 100X Coupler 1 unit 100X NAD+ 1 unit 100X Reagent Dye 1 unit 10X Blocking Buffer 1 x 6ml 10X Buffer 1 x 25ml 1X LDH-B Control 1 x 1µg Extraction Buffer (ab260490) 1 x 15ml LDH-B Microplate 1 unit Reaction Base Buffer 1 x 10ml -
Research areas
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Pathway
Fermentation; pyruvate fermentation to lactate; (S)-lactate from pyruvate: step 1/1. -
Involvement in disease
Note=Defects in LDHB result in deficiency of lactate dehydrogenase, a condition with no clear symptomatic consequences. Lactate dehydrogenase deficiency can probably be considered a non-disease. -
Sequence similarities
Belongs to the LDH/MDH superfamily. LDH family. -
Cellular localization
Cytoplasm. - Information by UniProt
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Alternative names
- Epididymis secretory protein Li 281
- HEL S 281
- L lactate dehydrogenase B chain
see all -
Database links
- Entrez Gene: 281275 Cow
- Entrez Gene: 3945 Human
- Omim: 150100 Human
- SwissProt: Q5E9B1 Cow
- SwissProt: P07195 Human
- Unigene: 446149 Human
Images
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Example control standard curve
Example control standard curve generaated using the method described in the protocl booklet.
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Lysate example curveLDHB activity was measured from Hela cell lysate, which contains hetero-tetramer isozyme composed of both (M) LDHA and (H) LDHB subunits. However, the liver derived cell line, HepG2, which contains mainly homo-tetramer of subunit M (LDHA), shows no enzymatic activity. -
Species cross reactivity shown with seraThe species cross reactivity of this assay was demonstrated by testing the sera from multiple species. The kit shows a good reaction with human, goat and bovine samples, but shows no reaction with mouse or rat sera which may also contain LDHB. This species cross reaction specificity is due to the species specificity of the capture antibody. -
Isoform specificity validated by Western blotLane 1: HeLa cell lysate
Lane 2: HepG2 cell lysate
Lane 3: LDHA (purified from rabbit skeletal muscle)
Lane 4: LDHB (recombinant)
Lane 5: HeLa – In-well extracted with capture mAb
Lane 6: HepG2 – In-well extracted with capture mAb
Lane 7: LDHA – In-well extracted with a capture mAb
Lane 8: LDHB – In-well extracted with capture mAbThe western blot results indicate that the capture antibody isolates active LDHB, but does not isolate LDHA (which would be detected by the anti-LDH antibody).