MitoSciences: MSFX-11 FlexPlex™ Module: Mitochondrial trifunctional protein (TFP) subunit alpha and beta (HADHA / HADHB)

Enzymes > Mitochondrial trifunctional protein (TFP) (HADHA / HADHB) > FlexPlex™ Module: Mitochondrial trifunctional protein (TFP) subunit alpha and beta (HADHA / HADHB)

FlexPlex™ Module: Mitochondrial trifunctional protein (TFP) subunit

alpha and beta (HADHA / HADHB)

Catalog No. MSFX-11

$125.00 - 1 module

The Mitochondrial trifunctional protein (TFP) subunit alpha and beta (HADHA / HADHB) FlexPlex™ Module is used to determine the quantity of TFP (mitochondrial trifunctional protein, subunit alpha / subunit beta) enzyme in a sample. A functional fully assembled TFP is immunocaptured within the wells by an antibody specific to the alpha subunit. The quantity of protein is measured by adding a second TFP specific antibody that is then labeled with horseradish peroxidase. This peroxidase changes the substrate from colorless to blue. The rate of color development is proportional to the amount of protein captured in the well and can be monitored at 600 nm. Alternatively, the assay can be terminated, at a user defined time, by the addition of 1N HCl (not supplied) and the assay performed as an end point measurement at 450 nm.

Click here to learn more about Mitochondrial trifunctional protein (TFP) subunit alpha and beta (HADHA / HADHB).

Specifications

Data

Documentation

Citations

Product Specifications
Applications:	analysis of TFP (HADHA / HADHB) quantity
Species Reactivity:	human
Range (min. 2x background):	5 - 40 µg / well (Example: HepG2)
Av. Intra-assay %CV (n=6):	15.8%
Storage Conditions:	Store at 4°C.
Country of Origin:	USA

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Specifications

Data

Documentation

Citations

Figure 1

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Figure 1. HepG2 titration series. Increasing amounts of HepG2 cells were analyzed using MSFX-11 to demonstrate the potential working range of the assay.

Figure 2

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Figure 2. HL60 titration series. Increasing amounts of HL60 were analyzed using MSFX-11 to demonstrate the potential working range of the assay.

Figure 3

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Figure 3. Human heart tissue titration series. Increasing amounts of human heart tissue homogenate were analyzed using MSFX-9 to demonstrate the potential working range of the assay.

Figure 4

	Fibroblasts extract loaded per well	Signal / background (control)	Reproducibility (%CV 6 wells)
MSFX-9 [MCAD]	40 µg	14	4.4
MSFX-10 [SCHAD]	10 µg	224	2.3
MSFX-11 [TFP]	1.25 µg	23	3.0
MSFX-12 [SOD2]	5 µg	4.8	12.2
MSFX-15 [PDH]	5 µg	4.3	21
MSFX-6 [GAPDH]	3.125 µg	86	2.7

Figure 4. The Trifunctional Protein (TFP) FlexPlex™ Module (MSFX-11) is designed to analyze the relative levels of TFP between samples. An experiment to demonstrate this is shown below in which fibroblast cell lines derived from two different patients with MCAD deficiency (MCADD) genetic disease are compared to a control fibroblast cell line (newborn human dermal fibroblast) and to a cell line with a deficiency in TFP.

Figure 5

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Figure 5. The raw signals can be compared between samples or in the example shown above the signals can be normalized to GAPDH. After normalization the patient cell line 1 has 93% reduction in MCAD and patient cell line 2 has a 92% reduction in MCAD. Both patient cell lines 1 and 2 are already described MCAD patients with the common mutation A985G. Interestingly, both patients have a greater than 50% increase in mitochondrial SOD2 expression. Patient cell line 3 has a normal expression level of MCAD but a 91% decrease in TFP enzyme. Patient cell line 3 is a cell line already identified as TFP deficient at the protein level but of unknown genetic origin. Interestingly, all three patient cells lines harboring mutations in MCAD or TFP have a 50% decrease in the level of SCHAD the short chain 3-hydroxyacyl-CoA dehydrogenase.

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