...led to their application for basic and translational research.  Basic research applications of scFvs: ...blood clearance, which is useful for imaging applications; better tissue penetration which is useful for... for therapeutic and imaging applications; and reduced immunogenicity when administered in vivo due to...from blood can be a drawback for therapeutic applications where longer retention times often increase ...laborious.  To avoid this, one or more of these approaches are often used for scFv expression in bacteria...since it’s genetically encoded.   Translational applications of scFvs: Just like antibodies, there are many...2019).Many of the scFvs designed for oncology applications are reformatted as bispecific molecules which...

...fits all When choosing an appropriate antibody first look at the application-specific data provided on...your specific application. Consider also that there may be variations within an application. Take immunohistochemistry... a linear antigen tend to work very well for applications that denature a protein to its primary (linear...while the website highlights a variation of your application, a publication successfully used it in a similar...plan to. To secondary or not to secondary?  The application will also determine whether you should use an... as it will help to amplify the signal. Some applications use either direct (primary-conjugated) or indirect... are secondary considerations to validation, application specificity, and experimental considerations ...

...your experiment (all well validated for your application, of course!) When the stars are aligned and the...factors that affect isotype choice. Target and application should be considered The first question you may... a strong signal and to work in a variety of applications. The desired specificity of your secondary antibody...affect my experiment?” Well, it depends on the application and target. If you are running a basic western...target is expressed at very low levels or your application requires a high degree of affinity, then species...considered when choosing an antibody. In some applications probing species-on-species, i.e. using a mouse...easily distinguishable from each other. In a direct approach, that is simple - but what about when you need...

...many applications, it is the only comprehensive method available. Targeted sequencing approaches More...differently. For a comprehensive approach, RNA-seq would be an appropriate method for an activator or repressor...lower them even further. While not all of these approaches will work for every experiment, at least one ...where only localization of the Cas occurs. 2 gRNA approach Off-targets that come from mutations arise from...More targeted alternative sequencing approaches have been developed. Cas ChIP-Seq, where only DNA sites ...especially when DNA mismatches are involved. Two other approaches have been used - GUIDE-Seq and integrase-defective...site-specific detection downstream. The con to these approaches is that NHEJ must perform the repair to allow...

... to be placed immediately upon request approval. Once approved, the request will remain in your account... in this blog post.     When will the expansion happen?  You will be able to request Packaged on Request...request will be reviewed by Addgene for legal approvals and technical compatibility with our production...process will take 1–2 days, and we anticipate approving the vast majority of requests.  If the vector ... request process.  Ordering If your request is approved, you will receive an email notification that your...is ready to order. An “Add to Cart” button will appear with the request in your Addgene account.  Orders...viral preps will be available at launch.  What happens after I order?  Once an order is placed, the Addgene...

...distinction of neighboring cells in imaging applications and the mapping of neuronal projections to their associated...while minimizing confounding secondary effects? An approach described as “knocksideways” (a British idiom ...domains, such as FKBP and FRB. The cell does not appear to mind extraneous proteins bound to its outer ...protein from expression to degradation. Chemical approaches may require expensive peptides or small molecules...and Technology Austria, decided to take a novel approach to control for these effects, drawing on the rapidly...optogenetics. Using a rational protein engineering approach, they designed Opto-RTKs, which activate signaling... on exposure to low-intensity blue light. This approach relies on the incorporation of light-oxygen-voltage...

... can happen at the national level within the executive branch or in Congress, or can also happen at the... there will be posts about virtual meet ups and happy hours that you can attend to help build relationships...arriving here in DC, is that so much policy actually happens outside of the government itself. You can work ...The “little p” policy is non-governmental work happening at large organizations who have to develop policies...who decides how they are going pursue regulatory approval for a new product or a well-endowed philanthropy...Program to learn more about where science policy happens.  When do you do science policy? Many science ...Project for some example proposals and consider applying for their accelerator for some hands-on guidance...

...library. Apply pooled library to cells. DIVERGE: Positive and negative screens POSITIVE screen: Apply library...control sample (no selection) In parallel, apply library Apply selection; most cells will survive Perform..., semi-random sequences that can be used for applications like lineage tracing or parsing the effects ...library Apply selection Most cells die or do not pass selection (in the case of a reporter) Sort “winning...genes involved in the selection NEGATIVE screen: Apply library Perform Next Generation Sequencing (NGS)...and control cells Generate a list of gRNAs that disappear with addition of the selection mechanism Positive...

...sense and appreciate this – and they will be happy too. On the flip side, if you are not happy at work,... If you are happy at work, you will find balance. If you enjoy working and come home happy in the evening...actions we can take to help ourselves can be an appropriate first step. Check out Joanne's Reddit AMA Managing...turns going; that way you can have someone there clapping more often than not. You want your family and ...tools and tactics to plan your work and make this happen. Be realistic and: Your co-workers will learn ... about their safety when they leave home which happens very, very fast! I will never forget the time my...figure out a way to keep being ourselves and to be happy. Building your plan to live a good life may be the...

... are approximately 4.1 kb in length, allowing them to be packaged in AAV.   Split AAV approaches  While...germline genome editing, but in vivo gene editing approaches have been limited by problems with Cas9 delivery...packaging capacity of AAV (~4.7 kb). While this approach has been proven feasible, it leaves little room...potent Cas9 protein, the Zhang lab took a different approach. They analyzed over 600 Cas9 orthologs and found...be divided into two groups: longer orthologs approximately 1350 amino acids in size, which includes SpCas9...SpCas9, and shortern orthologs approximately 1000 amino acids in size. From the pool of shorter orthologs...effects of SpCas9 and SaCas9, Zhang’s group used an approach called BLESS (direct in situ breaks labeling, ...

...platform offers a promising approach for experimental therapies and medical applications.  Find DIRECTED plasmids...Figure 1: Choosing the best PE6 variant for your application. Image reused form Doman and Pandey et al., 2023...REcipient Cells Through Envelope Design). This modular approach combines an interchangeable set of fusogenic and... CreVLPs, and eVLPs for in vivo and in vitro applications.   Figure 2: Modular features of DIRECTED...interactions or map a local proteome, but some applications have been challenging or inaccessible. The Ting...  Qin, W., Cheah, J.S., et al. (2023). Dynamic mapping of proteome trafficking within and between living... from different promoters, specific time points appear as fluorescent bands in the fiber, resembling a...

...fluorescence can be visualised. Another approach was to apply the same concept to lateral flow strips ... Suresh from Iowa State University. Since its appearance, SARS-CoV-2 has spread to almost every part of... required: 37°C heat block and 62°C heat block Approach: Viral RNA is extracted and subject to isothermal... required: 37°C water bath and 42°C water bath Approach: RNA is extracted from a sample and subject to...cleaved. Cleavage results in fluorescence and this approach can be incorporated into a lateral-flow readout...blue light illuminator or UV light illuminator. Approach: Highly similar to the SHERLOCK diagnostic test...microliter Time: Not mentioned in the paper, likely approximately 1-2 hours Equipment needed: Mini PCR machines...

...is an application in which antibodies that have been joined to fluorescent molecules are applied to a ...immunofluorescence There are a couple different approaches to be familiar with when talking about IF, direct...BioRender.com   Indirect IF is a much more common approach than direct IF, so we will focus on this technique... Primary Antibody This is the first antibody applied to your sample, which will bind to your target ...Unsurprisingly, this is the second antibody you apply, which binds to the first antibody and is conjugated...distribute antibodies validate them in a variety of applications, including IF (or IHC or ICC), and will share... time to research antibodies, protocols, and appropriate controls and you will be well on your way to ...

...sample of your protein of interest (POI). This approach is reliable, but it’s overkill in many cases. ...than “condition 2.” By eye, the loading control appears about the same between the two conditions. But ...detected with an antibodies, just like the POI. This approach is so common that we’ve used it for our cartoon...cells. This means that at sample concentrations appropriate to detect a less-abundant POI, the loading control... control aren’t too far apart, they may have overlapping linear detection ranges, allowing you to accurately..., 2008; Gilda & Gomes, 2013). Depending on the approach you choose, your loading control measurement will...on the same gel or membrane (depending on your approach) as your final blot. Quantifying the entire lane...

...Blocked by overlapping methylation Not affected BsaI Not affected Blocked by overlapping methylation...Blocked by overlapping methylation HpaI Not affected Not affected Blocked by overlapping methylation...Blocked by overlapping methylation Not affected Not affected MscI Not affected Blocked by overlapping methylation...Not affected Blocked by overlapping methylation XbaI Blocked by overlapping methylation Not affected...methylation Not affected ClaI Blocked by overlapping methylation Not affected Not affected DraI Not affected...

...think “but that won’t happen to me, I have a backbone”. The sad truth is, if it happened before, it almost...choosing a lab, but almost all of these guidelines can apply to any workplace or job. It amazes me when the most...scheduled or only ad hoc? Does the mentor keep appointments with group members or are they frequently cancelled...conflicts in the lab? Do they motivate with the appropriate balance of encouragement and criticism? Do they...was told that the PI removed a woman from the appropriate first authorship position on a publication of...and profoundly regretted it when the same thing happened to her a year later. I hear too many science trainees...almost certainly will also happen to you. As you speak to lab members and alumni, don’t ever disregard warning...

...t just happen to you or me (though, let's be clear, they have all happened to me.) They happen to everyone...is, when it happens to other people, it’s normal and quickly forgotten, and when it happens to you, it’...communicator, I’ve done a lot of public speaking, and I'm happy to share what I've learned along the way. Own your...comes to public speaking, Murphy’s law definitely applies: what can go wrong, will go wrong. The trash removal...interest incorrectly on slides 8-16. Your notes disappear. The wrong version of your slide deck gets uploaded...completely normal to be anxious, nervous, and/or apprehensive. I still get nervous quite regularly, and I'...

...AID an appealing tool for studying essential proteins. Until recently, it was challenging to apply the AID-tagging...brief articles highlight the main features and applications of a partiular plasmid or set of plasmids. We...December 2016 (Q3) MARCH 2016Enhancer activity mapping with STARR-seq   Although many technologies have...classical TANGO assay, Kroeze et al. developed a new approach termed Parallel Receptor-ome Expression and Screening... (PRESTO-TANGO) to screen the NCC-1 library of approved drugs against the entire kit; a parallel analysis...multiple expression systems before they achieve the appropriate expression level for their gene of interest. ...GMAP-compatible vectors for a wide variety of biological applications. First, GMAP-compatible backbones for lentivirus...

... drug approval boards are being finalized, and it is very likely the treatments will be approved for use...disease. Meanwhile, Hans Clevers and colleagues applied CRISPR to disease correction in adult stem cells...CRISPR-based editing therapies that are currently approved or have made significant progress in clinical ...Therapies Cancer and CAR-T therapy The FDA has already approved CAR-T immunotherapy for cancer treatment by traditional...These therapies are delivered with an autologous approach, editing patient cells ex vivo and then returning...but especially concerning when using a systemic approach. Regarding effectiveness, if the editing frequency...meaningful way. Delivery, especially for systemic approaches, has its own unique set of obstacles. Thus far...

...which subpopulation of DA neurons could activate appetitive rather than aversive stimuli. In a recent paper...rAAV FLEx-based vectors to answer this question, mapping and characterizing the activity of the DA neurons...pursue a teaching career, and she now is a very happy scientific communication manager at genOway.     ... cell types for imaging and long-range circuit mapping." Journal of Neuroscience 28.28 (2008): 7025-7030...Philippe Coulon, and Ilka Diester. "Optogenetic approaches to study the mammalian brain." Current opinion...Yizhar, and Karl Deisseroth. "The development and application of optogenetics." Annual review of neuroscience...Mei, Yuan, and Feng Zhang. "Molecular tools and approaches for optogenetics." Biological psychiatry 71.12...