April 2021 News

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      We publish another collaboration between scientists at the University of Florida and EnCor, “TNF increases tyrosine hydroxylase expression in human monocytes” by Madison et al., BioRΧiv 10.1101/2021.04.13.439627v1, It describes in detail an ELISA developed using EnCor antibodies which can be used for quantitation of tyrosine hydroxylase (TH) levels in a variety of experimental and clinical contexts. The specific antibodies used are our mouse monoclonal antibody MCA-4H2 and our rabbit polyclonal RPCA-TH. They also used our chicken polyclonal to the cell division marker Ki67 CPCA-Ki67. Previously the same group showed that TH levels were detectable in blood cells of healthy control and Parkinson’s patients and that the levels detected in Parkinson’s patients were significantly higher, a potential very significant finding.
      We have been very interested in recent studies of NF-L, neurofilament light, as a biomarker of neuronal damage and degeneration. Much of the recent work has been performed with the excellent Uman NF-Light Assay. The epitopes recognized by the two antibodies used in this assay are not known, and so we have been working to fully characterize them. As part of this work we generated a codon optimized cDNA designed to express amino acids 256-400 of the α-helical rod region of human neurofilament NF-L, specifically the Coil 2a and Coil 2b regions. Our full length recombinant human NF-L protein Prot-r-NF-L is widely used as a protein standard in ELISA, Simoa and other kinds of antibody based assays for NF-L detection, including the Uman assay. Our new truncated product, Prot-r-NF-L-rct, contains the epitopes for both the capture and detect NF-L monoclonal antibodies used in the Uman NF-Light assay and also the epitope for our mouse NF-L monoclonal MCA-1B11. This product may therefore be a more convenient protein standard for use with the Uman and similar NF-L assays, and our new data suggests that MCA-1B11 may form the basis of further NF-L assays.
      We also updated our product data for several of our recombinant proteins including mCherry Prot-r-mCherry and GFP Prot-r-acGFP.

February 2021 News

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      We published a preprint on the BioRΧiv server, observations on the biomarkers pNF-H and UCHL1 in the bodily fluids of patients recovering from a common type of thoracic surgery. This report “A study of the neuronal injury biomarkers pNF-H and UCHL1 in serum, CSF and urine in a cohort of thoracic endovascular aortic repair (TEVAR) patients”, by Beck et al., BioRΧiv 10.1101/2021.03.02.433405v1, in addition to documenting blood, CSF and urine levels of these two proteins in a large cohort of TEVAR patients, also provides important quantitative data on the levels of UCHL1 in healthy control blood samples using multiple antibodies, results with antibody data confirmed by mass spectroscopy. We also document the presence of UCHL1 in certain urine samples. We used novel pNF-H and UCHL1 assays developed for the MSD electrochemiluminescent platform using EnCor antibodies and ran all the several hundred assays in the EnCor lab. We also include epitope mapping and other data on our panel of UCHL1 antibodies including the widely used mouse monoclonal antibody MCA-BH7.
      We have updated product data for our recombinant full length human proteins, NF-L, GFAP, Vimentin and UCHL1, products PROT-r-NF-L, PROT-r-GFAP, PROT-r-Vim and PROT-r-UCHL1 respectively. We now provide the exact amino acid sequence and composition, isoelectric point, molecular weight, extinction coefficient and other relevant data for each protein. These proteins are widely used as protein standard for ELISA, MSD, Luminex and Simoa type assays, and also as immunogens for antibody production. We will provide this information for all of our recombinant proteins shortly and have already done this for our full length recombinant rat GFAP protein PROT-r-GFAP-rat.

January 2021 News

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      We release novel antibodies to one of the CasΦ, a.k.a. Cas-12j, family of enzymes. These are newly described and exceptionally compact enzymes of great potential utility in gene editing by CRISPR. Recent ecosystem and microbiome DNA sequencing and assembly characterized numerous whole genomes of “huge phages” or megaphages, see Al-Shayeb et al. Nature 578:425-431 (2020). These phages have much larger genomes than those of well studies bacteriophages such as phage-λ and members of this family are apparently ubiquitously found in all microbiomes and ecosystems. Interestingly the genomes may include genes which encode novel CRISPR associated or Cas enzymes. The lab of recent Nobel prize winner Professor Jennifer Doudna further characterized some of these megaphage Cas enzymes and showed that they could be used to perform CRISPR but had the significant advantage that they were much smaller in molecular size than typical Cas9 and Cas12 enzymes such as those from S. pyogenes and S. aureus, see Pausch et al. Science 369:333-337 (2020). The amino acid sequence of several of these enzymes was published and the group studied was named CasΦ or Cas12j. The small relative size of CasΦ enzymes leaves more room for other nucleic acid sequences in AAV and other viral vectors which typically have a limited DNA capacity. We made a mouse monoclonal and a rabbit polyclonal to one of these enzymes, CasΦ-2, MCA-5F95 and RPCA-Cas12J respectively. These can be used verify correct expression of CasΦ-2 protein in CRISPR experiments.
      We have also made a novel, high quality and very high titer rabbit polyclonal antibody to microtubule associated protein 2 (MAP2), RPCA-MAP2. Antibodies to MAP2 are excellent markers of neuronal dendrites and perikarya and are also useful in developmental studies. This new antibody was made against recombinant forms of the entire MAP2 projection domain using a mixture of our three human MAP2 projection domain constructs and works very well on tissues of transgenic and wild type rodents at dilutions of 1:10,000. It is therefore a very cost effective reagent for routine neuronal identification in cell cultures and tissue sections.

December 2020 News

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      We add new data concerning our three SARS-CoV2 spike protein cell binding domain antibodies which, we now show, strongly bind to Vero cells infected with patient derived SARS-CoV2 virus. Vero cells are a cell line derived from monkey kidney which express a membrane localized ACE2 receptor very similar to the human form and are therefore susceptible to SARS-CoV2 spike protein binding, internalization and infection. Infected and control Vero cells were grown in culture, fixed, and stained with the DNA dye DAPI, revealing cell nuclei. Cells were then reacted with EnCor antibodies to the cell binding domain of the SARS-CoV2 spike protein, namely MCA-2G1, MCA-5G8 and RPCA-SARS-CoV2-bd. The virus could clearly be seen to accumulate in the cytoplasm of infected cells, press here to see the relevant image.
      We also continue to experiment with our confocal microscope and can now reliably produce four color images making use of our growing collection of antibodies made in different animal species. For example we just deposited a four color image of HeLa cells to Wikipedia. This image was made using our mouse monoclonal to β-tubulin MCA-1B12 in green, rabbit antibody to HSP60 RPCA-HSP60 in yellow, chicken antibody to fibrillarin CPCA-Fib in red and with the DAPI DNA stain in magenta. The β-tubulin antibody reveals the microtubular cytoskeleton, the HSP60 reveals mitochondria, the fibrillarin antibody reveals nucleoli and the DAPI reveals cell nuclei. A link to this Wikipedia image is here.
      We also continue to add to the characterization of our monoclonal antibodies with more epitope data on our mouse monoclonal antibody to neurofilament NF-L MCA-1B11. The epitope is dependent on a short peptide at the beginning of the “Coil 2” region now known, by analogy to the structure of the related protein vimentin, to correspond to a continuous almost straight α-helix. Interestingly the two monoclonal antibodies to NF-L used in the Uman NF-light Assay and also in the Quanterix Simoa NF-light Advantage Assay both bind to this same small region. We will provide more specific information on these findings shortly and will likely publish a detailed scientific report on these and other findings we have made on widely available NF-L antibodies.
      Finally we open an Etsy store, see here! So far we only listed our brain cell wrapping paper but other products will be added in the near future. Oh, and best wishes for a safe and Happy Holiday Season!!

August 2020 News

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     We add another mouse monoclonal antibody to the astrocytic and biomarker protein GFAP, MCA-3E10. This antibody was made against recombinant human GFAP and the epitope and kinetics of this antibody are known. It has utility as a component of assays designed to detect GFAP in blood, plasma and sera, which has become of potential interest as a surrogate marker of CNS damage and disease. We also release a chicken polyclonal antibody to neuron specific enolase, a.k.a. enolase 2, CPCA-NSE and a rabbit antibody to fatty acid binding protein 7, RPCA-FABP7. Finally we market two new antibodies to annexin A6, a useful marker of cellular membrane damage and “blebs”, herniations of the cell membrane associated with apoptosis. These are mouse monoclonal MCA-4G3 and rabbit polyclonal RPCA-ANXA6.

July 2020 News

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      Like many other laboratories with expertise with antibody production we have focused some urgent attention on the SARS-Cov2 virus and the ACE2 protein which it uses to enter and infect human cells. The region of the virus spike or S-protein and the corresponding virus binding region on the ACE2 protein were recently characterized so we made recombinant forms of both regions in E. coli (Prot-r-SARS-CoV2-bd and Prot-r-ACE2-bd). We now have a panel of excellent mouse monoclonal antibodies to the ACE2 binding site of the SARS-CoV2 S-protein, and are marketing the first two of these, MCA-5G8 and MCA-2G1. We also made an affinity purified rabbit polyclonal antibody raised against the same immunogen, RPCA-SARS-CoV2-bd. These antibodies can be used to identify the SARS-Cov2 virus on western blots and by immunofluorescence and may also block the binding of the virus to ACE2. Of general interest to the world, the SARS-CoV2 S-protein ACE2 binding region turns out to be an excellent immunogen, so that it is likely an ideal target for vaccine development.
      We release new antibodies to β-synuclein, a mouse monoclonal and a rabbit polyclonal, MCA-6A10 and RPCA-SNCB. β-synuclein, like the closely related α-synuclein, is a major brain protein concentrated in synaptic regions. We also release new antibodies to Annexin A5 MCA-6A12 and RPCA-ANXA5. Annexin A5 is a Calcium and membrane associated protein which is upregulated when cells become apoptotic. We also release two new mouse monoclonals directed against PEA-15 MCA-4D2 and MCA-4D117. This protein was discovered as a “phosphoprotein enriched in astrocytes of 15kDa” (PEA-15) and independently as a “protein enriched in diabetes” (PED) and so is sometimes referred to as PED/PEA-15. PEA-15 is one of the large family of proteins containing a death effector domain (DED) and appears to have important functions in the regulation of cell growth, apoptosis and glucose metabolism. Finally we release a goat polyclonal antibody to myelin basic protein, GPCA-MBP, which can be used to identify and study myelin sheathes, oligodendrocytes and certain types of Schwann cell.

June 2020 News

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      We add more goat antibodies to high value targets, namely to 2′-5′-cyclic nucleotide phosphodiesterase (CNP), GPCA-CNP, glial fibrillary acidic protein (GFAP) GPCA-GFAP, and mCherry GPCA-mCherry. Antibodies to CNP are excellent markers of myelin sheathes, GFAP antibodies label astrocytes, neural stem cells and a few other nervous system cells while mCherry is a widely used genetically encoded fluorescent tracer. Also another scientific paper performed using EnCor antibodies with the described assays run in the EnCor Lab, Holmström et al. 2020. This paper measures the levels of phosphorylated neurofilament heavy chain (pNF-H), ubiquitin C-terminal hydrolase 1 (UCHL1) and GFAP in the cerebrospinal fluid (CSF) and sera of recovering spinal cord injury patients and concludes that pNF-H and GFAP levels particularly in CSF may be useful for future monitoring of damage, secondary injury and response to treatment in these patients.

May 2020 News

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      As every other company we have been affected over the last couple of months by the Covid-19 situation. However we continue to do essential work with reduced hours and remain open for business. We have been shipping to our various OEM partners as before, and, since the receiving departments of many university, company and research institutes are closed down we can and have been shipping to researchers home addresses. We have also initiated some SARS-CoV-2 related research which will result in some new products, including recombinant forms of the cell binding domain of the major coat protein of the SARS-CoV-2 virus PROT-SARS-CoV2-bd and the extracellular domain of the angiotensin converting enzyme 2 (ACE2), now known to be where the SARS-CoV-1 and SARS-CoV-2 viruses bind to the surface of human cells, PROT-ACE2-bd. This binding interaction is essential for virus internalization and infectivity so that antibodies and other reagents that interfere with this specific interface are of great potential clinical interest. We also add more data on the properties and utility of our older products. For example we have epitope mapped our widely used monoclonal antibody to UCHL1 MCA-BH7 and now show that our chicken antibody to tyrosine hydroxylase CPCA-TH can be used to isolate TH positive PBMCs by fluorescent cell sorting. For details of this see Gopinath et al. (2020), another EnCor/academia collaborative study.

February 2020 News

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      We release a new antibody to FOX3/NeuN, GPCA-FOX3, an excellent marker of neuronal nuclei and perikarya. The original NeuN antibody has been widely used to identify and count neurons since only these cells express this protein. The NeuN immunogen was found to correspond to the RNA binding protein FOX3, and our new antibody was made against a recombinant construct including the N-terminal 100 amino acids of human FOX3. The antibody was made in goat and so is very useful for double and triple label of mouse and other rodent species.

January 2020 News

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January 2020 News

      Happy New Year! Although EnCor is primarily a commercial enterprise we still do have several ongoing interesting collaborations with academia. For instance we were involved in a recent high profile publication in molecular archaeology; a severed human head was found in surprisingly well preserved state in a pit in Heslington, near York, England and dated to 2,600 years ago. The original owner of the head had presumably been executed or sacrificed, something which apparently happened quite a lot in pre-Roman Britain. This skull was found to contain a shrunken but surprisingly well preserved brain, which generated considerable interest, see the Wikipedia page here. EnCor in collaboration with a group of European scientists showed that tissue from this brain was able to elicit immune response to two important brain proteins, GFAP and MBP, indicating that these proteins were surprisingly stable and still immunogenic over more than two and a half millennia. These findings were confirmed using mass spectroscopy and other experiments showed that many other brain proteins were partially preserved. The paper was just published (see here) in the Journal of the Royal Society Interface. The report has generated considerable media interest, see for example here, here and here, or just do a Google search for “2,600 year old brain”.