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Available ordering format
Liquid in PBS pH 8.0, 0.02% sodium azide
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Immunogen
KLH-conjugated synthetic peptide derived from known bacterial NifH subunits of bacterial nitrogenase enzymes of the FeMoCo type including Synechoccocus sp. Q2JP78 , Trichodesmium theibautii, Anabaena sp. P33178 and Nostoc sp. Q51296
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Raised in
Hen
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Clonality
Polyclonal
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Clone
Polyclonal
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Purification
Affinity purified IgY
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How to reconstitute
See included datasheet or contact our support service
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Storage condition
store at 4°C; make aliquots to avoid working with a stock. Please, remember to spin tubes briefly prior to opening them to avoid any losses that might occur from liquid material adhering to the cap or sides of the tubes.
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Verified applications
immunohistochemistry (IHC), immunofluorescence (IF),western blot (WB)
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Connected products
AS01 021S | NifH protein standard for a quantitative western blot Recommended secondary antibody for chemiluminescent detection, AS10 1489Recommended secondary antibody for other detection systemsAlgal protein extraction bufferSecondary antibodies
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Recommended dilutions for use
1: 2 000 (WB), 1: 500 (IHC), 6 µg/ml (IF)
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Molecular weight expected аpparent
27 | 32.5 kDa
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Verified reactivity
Anabaena PCC7120, Clostridium butyricum, Codakia orbicularis, Cylindrospermopsis raciborskii CS-505, Nostoc sp, Rhodopseudomonas palustris, Trichodesmium sp., nodules of Trifolium repens L.
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Possible reactivity
Desulfotomaculum reducens (strain MI-1),Clostridium cellobioparum, Magnetococcus sp., Methanobacterium thermoautotrophicum, Methanococcus maripaludis, Methylobacterium sp , Rhodopseudomonas palustris TIE-1 strain, alpha,gamma,beta proteobacteria, enterobacteria, low GC gram+, high GC gram +, euryachaeotes, Azotobacter vinelandii (Gram-), Klebsiella pneumonia, cyanobacteria, Enterobacter genera, able to fix atmoshperic nitrogen, Rhizobium meliloti
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No reactivity
Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803 as NifH protein is not present in those cyanobacterial species, Frankia sp.
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Supplementary information
An enzyme involved in chlorophyll synthesis, present in all cyanobacteria (fixing and non-nitrogen fixing) is a member of the NifH family/superfamily. Agrisera anti-NifH antibody will not show a strong reactivity to this target.In photobionts like Anabaena sp., low nitrate growth is required to turn on the NifH expression to high enough levels to detect NifH protein. Immunofluorescence protocol Insect dissected tissues (digestive tract, fat body, carrying NifH positive bacteria) of large workers were fixed in cold methanol (20 min, -20°C) and then permeabilized in cold acetone (5 min, -20°C). Samples were subsequently rinsed three times with PBS with 0.1 % Triton-X 100 at RT (PBST) and incubated for 5 minutes in PBST. This was followed by incubation of tissues for 1 hr with 6 ug/ml affinity purified anti-NifH antibody (Agrisera, AS01 021A) diluted in PBS-TBSA (PBS, 0.1 % v/v Triton-X-100, 1 mg/ml BSA) and 3 washings with PBST. Samples were then incubated in the dark with a goat anti-chicken IgY conjugated to Dylight 488 (Pierce, SA5-10070) for 45 min and were washed twice (PBS, 0.1%v/v Triton-X-100). Finally, the tissues were mounted in Vectashield medium containing DAPI (Vector Laboratories, H-1500) and viewed under a SP5 Leica confocal microscope with 10X and 63X objectives. Courtesy of Drs. Panagiotis Sapountzis and Mariya Zhukova, University of Copenhagen, Danmark
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References
König et al. (2016). Nitrogen fixation in a chemoautotrophic lucinid symbiosis. Nat Microbiol. 2016 Oct 24;2:16193. doi: 10.1038/nmicrobiol.2016.193. Liberti et al. (2015). Bacterial symbiont sharing in Megalomyrmex social parasites and their fungus-growing ant hosts. Mol Ecol. 2015 Apr 24. doi: 10.1111/mec.13216. Calusinska et al. (2015). Genome-wide transcriptional analysis suggests hydrogenase- and nitrogenase-mediated hydrogen production in Clostridium butyricum CWBI 1009. Biotechnology for Biofuels (2015) 8:27. Moirangthem et al. (2014). A high constitutive catalase activity confers resistance to methyl viologen-promoted oxidative stress in a mutant of the cyanobacterium Nostoc punctiforme ATCC 29133. Appl Microbiol Biotechnol. 2014 Jan 3. Chen et al. (2013). Improving conversion efficiency of solar energy to electricity in cyanobacterial PEMFC by high levels of photo-H2 production. Int J of Hydrogen Energy (2013):1-8.(Anabaena, western blot)Plominsky et al. (2013). Dinitrogen Fixation Is Restricted to the Terminal Heterocysts in the Invasive Cyanobacterium Cylindrospermopsis raciborskii CS-505. PLOS ONE, Open Access.
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Scientific context
Nitrogenase is involved in biological fixation of atmospheric nitrogen to ammonia. Alternative protein names: nitrogenase component II, nitrogenase Fe protein, nitrogenase reductase, FeMoCo-nitrogenase.
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Notes
antibody concentration is 1.36 µg/µl
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Protein number
Q2JP78 , P26254 , P33178 , Q51296
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TAIR number
Refer to NCBI
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