Mono-a-MonoClonal: A Biotech Blog
SBIR: Thrill and Agony
Whenever one thinks about small business innovation research grants (SBIRs), you can’t help but recall ABC’s Wide World of Sports quote, “The thrill of victory. The agony of defeat.” Both SBIR grant writers and SBIR reviewers experience the highs and lows on both sides of the sport, so to speak.
A tremendous amount of effort goes into submitting an SBIR application: months of generating preliminary data, writing, re-writing, obtaining support letters, justifying budgets, and so on. Then you hit the submit button, hope for no errors, and wait. And wait. And wait. With luck, you get a good score and a positive summary statement from your scientific review group. And then you wait again as the application sits in the parent institute awaiting the green light. Many of NeoClone’s grants go through the National Cancer Institute (NCI) which does not publish pay lines, so it’s always a guessing game if the program officer will like the grant and advocate for it, or if it will go unfunded (and undergo subsequent revision and another submission). For example, we submitted a grant August 5, 2011 and it funded over a year later, September 1, 2012 One could say there is an additional agony of waiting, as well as of defeat. The thrill comes when you get a good score, a great review from the study section (scientific review group/SRG), and when the application funding comes through. There is nothing like the relief, and the pride, you feel at each of those 3 stages.
On the other hand, for SBIR reviewers sitting on those SRGs, it’s less a marathon and more like a sprint in a crowded room. Despite popular opinion, most reviewers are dedicated to doing a good job and take their role as advocate for the grants they’re assigned very seriously. And YES, they have actually read the grants they’re assigned. It’s very exciting when a proposal you strongly advocate for gets a better score because you argued for it, and that is a great win to have. It’s also a thrill to be assigned a phase II application from investigators whose phase I application you previously reviewed and scored well. Conversely, it’s very disappointing when something you feel is a quality application scores worse after discussion despite your best efforts, or a lower quality grant gets a higher score than you felt it deserved. Overall, though, it’s great to be part of the process, and scientific innovation is the ultimate winner.
So here at NeoClone we are preparing to enter the grant submission marathon once again. We have to admit, we’ve never been huge fans of distance running.
Western Blot Woes- Part 3
What should you do when you need to repeat a western blot? Most of us have been in this position for a variety of reasons. The background was too high. The target signal was too faint or too strong. The transfer was incomplete. You might need a different percentage gel to provide clearer resolution. Or perhaps you will need to compare antibodies from various lots or providers over time. It’s quite frustrating and time-consuming to repeat the whole process again. The good news is there are several options available to minimize efforts associated with repeating western blots.
Save those samples. Denatured samples for SDS-PAGE and western blotting are typically very stable. Assuming lysates, media, sera, or other test material are not limiting, you might want to prepare additional sample volume for SDS-PAGE. If you typically load 10ul/lane, consider preparing 20, 30, or even 50ul of each sample, especially if this is your first time evaluating a new antibody against its target. The remaining sample is usually stable when refrigerated or frozen and in most instances the sample may be used directly without heating to denature again. This makes repeating the gel and transfer much easier.
Plan ahead and save un-probed blots. Whenever you are using only a partial gel and blot, consider loading all the wells. The portion of the blot that is not needed today can be cut away from the portion to be probed. The remaining section can then be dried after blocking and saved by storing it between two pieces of clean filter paper placed in an envelope or some similar storage method. You can delineate sets of lanes with pre-stained molecular weight markers for accurate cutting of the blot. You should also include a copy of the information detailing the lane contents, sample preparation, gel percentage and membrane type with the stored blot. Dried blots are typically very stable at ambient temperature.
Western Blot Woes- Part 2
What do you do when your western blot just isn’t working? The control samples that should contain detectable amounts of your protein aren’t providing positive results. The vendor’s western blot control isn’t detected either. Now you start asking questions like: “Is the primary antibody bad?” The technical support staff from the antibody vendor assures you that the antibody you received works fine in their hands. This can be a frustrating situation. The good news is that there are alternatives to consider.
If a careful repeat of the experiment produced the same negative results, the best advice is to try something different. You could try a radically different detection method, for example, but that can be expensive and time-consuming. You could evaluate various substrate buffers or membranes or secondary antibodies. Perhaps the easiest thing to do would be to contact the primary antibody manufacturer and ask how they use or test the antibody and then try to replicate their methods.
We discovered the importance of testing the basics while executing new long-term stability studies for several of NeoClone’s antibodies. Negative western blot results were observed for several antibodies that had performed spectacularly in the past. Multiple lots of these antibodies that were known to work well no longer yielded positive results in the new western blot study. Paradoxically, several other antibodies tested at the same time were still working fine.
A thorough investigation revealed that different methods were employed than were previously used for our standard Quality Control western blot testing. The method that was not working consistently utilized PVDF membranes and a diethanolamine-based alkaline phosphatase buffer drawn from the literature. Previous in-house QC western blots that worked consistently for all the antibodies being tested utilized nitrocellulose membranes and a tris-NaCl-magnesium chloride-based alkaline phosphatase buffer. When all the antibodies were re-tested using nitrocellulose and a tris-NaCl-magnesium chloride-based alkaline phosphatase buffer, positive results were obtained for all the antibodies.
We rediscovered that it is important to always compare methods, materials and reagents to previous experiments as well as published sources before starting any new procedure. Results in your lab may vary from the vendor’s experience due to legitimate variations in these and other factors.
Additional Thoughts on China
Although NeoClone has yet to close on a deal (commercial or investment) in China, given our current activity in the space, we remain cautiously optimistic that it is only a matter of time before this happens. So once again, I find myself getting lots of comments on ‘what to expect’ when dealing with the Chinese. And, once again, I find that conventional wisdom is not meshing with our reality. Here are a few more of those myths vs. reality….
Opinion #4: Chinese investors only want to invest money in China. We have found that this is simply not the case. Yes, there are many opportunities for investment in China, but they are also looking globally, just like everyone else. As NeoClone contemplates its next round of investment, we will be speaking to Chinese investors, and while it is impossible to predict the outcome, they have expressed interest in investing in our business ‘as is’ – which currently does not have any Chinese presence.
Opinion #5: The Chinese will want to change the deal at the ‘last minute’. Now, while we have not gotten this far with an investor or company, to date this has not been the case. We have not come across any circumstance where this has been the case. In fact, we have experienced the opposite, once we come to terms on a part of a deal, no one has suggested changing terms. This is not to say that terms have not been modified along the way; this occurs in the vast majority of deals we have done, but no ‘unexpected’ changes or requests.
Opinion #6: Traveling to China is difficult. While it is true that you do need a Visa, we did not find it particularly difficult (or expensive) to obtain, and they can be good for a year. Additionally, you can get flights out of most of the major US airports direct to Beijing or Shanghai, so aside from a long flight, it is not particularly difficult to travel to China.
As I said above, we do not currently have any completed business deals in China, but as the inevitable approaches, we get more and more comfortable that while it is a new market, it is not something to be feared, but embraced.
Reflections on BIO
So our CEO, Deven McGlenn, and our CSO, Uwe Muller, attended the BIO International Convention held in Boston recently. This was not their first time at this meeting, but even having some idea of what to expect, they continued to be impressed at the size (well over 16,000 attendees from 60+ countries) and scope (if its biotech related in any way, its there) of this meeting. To be clear, this is a meeting about business development, not science. People are not there to learn about new ground breaking research, they are there to learn about new and innovative products and services. It also has a partnering feature where you can request and schedule meetings with others at the convention, making the meeting far more productive. Additionally, Deven thinks the convention does an excellent job of creating interaction opportunities at many levels.
So after having 25+ formal meetings, attending over 15 different events and walking the very sizable convention exhibit area, one word kept coming to mind – networking. While they certainly found value in their more formal interactions, they would say the meeting was just as productive from just ‘bumping into’ others at the various events and on the exhibit floor. While it would be exceedingly difficult to quantify the ‘value’ of this meeting, they easily talked to over 10 of our existing clients (many from overseas), already have formal follow-ups with 7 new companies, and an additional ~ 15 good leads. We would have to classify the meeting as very valuable. However as this week set in, we are reminded that having all of these great opportunities means a whole lot of emails to everyone they saw last week! But before we go back to - “Dear Sirs, it was great to see you at BIO last week….” Deven and NeoClone would like to extend a formal thank you to BioForward (www.bioforward.org/) and the Wisconsin Economic Development Corporation (wedc.org/) for their financial support of NeoClone to attend this meeting. Now back to the part of the meeting requiring the most work, the follow-up!
Western Blot Woes Part 1.
Western blotting is a relatively simple and common technique to assess the presence or absence of a protein in an often heterogeneous sample. Following gel electrophoresis of the sample, proteins are transferred to a membrane and are probed by antibodies specific to the protein of interest and detected either colorimetrically or by chemiluminescence. There are many different protocols available to do this and vary by membrane type (PVDF vs nitrocellulose), buffers, secondary antibody-enzyme complexes (horse radish peroxidase or alkaline phosphatase), substrate and detection equipment.
Problems may arise when results are ambiguous, expected signals are absent, or additional signals and high background are observed. There are a number of factors that one might consider when trouble-shooting western blot problems.
If you have not previously used a new primary antibody, consider demonstrating that the secondary detection antibody recognizes the primary antibody. Include a lane of denatured primary antibody in the gel and blotting procedure. Many secondary antibodies react with both heavy and light chains. If the heavy and light chains of the primary antibody are observed, then the primary and secondary antibodies are compatible for western blotting. This control is also evidence that the blotting and detection procedures were properly executed.
Additionally, consider using a western blot control containing the target antigen if one is available for the primary antibody, possibly the recombinant protein that was used to create the primary antibody. Antibody manufacturers often use these proteins for Quality Control testing of the primary antibody and may be available in their catalog.
If you are unsure of the quality of your heterogeneous sample, then simultaneous probing for an internal control protein may also be a good control. Pick a protein that is known to be present in the sample and has been detected previously by western blot using the appropriate antibodies. Typically a single blot can be probed with multiple primary antibodies at their appropriate dilutions without issue, as long as the target antigens are of different sizes. The same could apply to secondary antibodies as long as the two primaries were produced in different hosts, say goat and mouse.
Look for more western blot tips soon.
From the Bench
There is usually more than one way to do a job. Most of us have heard the common idioms: “There’s the right way, the wrong way, and army way” and “There’s more than one way to skin a cat.” Most people also realize that if something isn’t working as expected it doesn’t make sense to keep trying the same thing repeatedly. Or as Albert Einstein said “Insanity is doing the same thing over and over again, but expecting different results”.
In biological research, all scientists have had these experiences. The assay isn’t working. The reaction isn’t producing the expected product. Experiences like this lead to frustration and of course, the ultimate question: “What am I doing wrong?”
Even when experiments are working well, scientists are always coming up with better and more efficient methods for laboratory procedures. Some of these improved techniques are so useful they become kits and are available to everyone. But many other alternative techniques are embodied as part of a highly experienced scientist’s ‘tricks of the trade’ and thus not widely available as useful information for others.
The good news is that NeoClone scientists are teaming up to share some of our collective knowledge regarding various laboratory techniques. Our hope is that the tips and technical advice we offer will be helpful if you encounter problems in the lab.
Look for our bench tips in this blog space in the coming weeks.
Reflections on China
We were recently given an excellent opportunity to introduce our antibody offerings (with a particular focus on our newly developed therapeutic capability) to the Chinese marketplace. Not having done much business in China prior to this, and this being my first trip to this spectacular country, I really did not know what to expect. That said, LOTS of other people had perspectives / opinions on what I should expect in trying to do business with Chinese companies – some of it was good advice, while some of it I suspect was more borne out of lore, rather than reality! So following are some of what our experiences actually were, vs. what ‘conventional wisdom’ might suggest we should have experienced…
Opinion #1: The Chinese will try to steal your technology. This was by far the most commonly repeated phrase we heard prior to our trip. While I am no expert on this topic, this is certainly a theme I have heard / read about many times and was of course aware of prior to our arrival. However, our experience was different, and I do not consider this a primary concern anymore. What we experienced, particularly as it relates to highly regulated products like therapeutic antibodies, was that the Chinese companies are just as concerned with getting their products through their version of the FDA approval process as US / European companies are in getting regulatory approval. It also became very clear to us that it is extremely unlikely that a product will gain regulatory approval in China if the intellectual property is in question. So while this “conventional wisdom” may be true in other industries, we do not see it as a factor when doing business there.
Opinion #2: It takes forever to get a deal done in China. Given this was the first time we were offering our therapeutic antibodies to the market in general, we did not have expectations of getting a ‘quick deal’. However, our experience would suggest that perhaps this conventional wisdom may not hold true. We met with about a dozen Chinese companies of various sizes and the reality was that the bigger the company, the slower the process was going to be, and depending on the real need, that would drive the timing of getting a deal done. This is our same experience everywhere else in the world! There does not seem to us to be any obvious reason why it would be inherently slower to do business in China. To the point, while we have not closed on a deal yet, and it is still possible we will not get it done, it is very possible that we will have our first therapeutic client before the end of Q2 this year!
Opinion #3: The Chinese market is HUGE! Our perspective in this area is mixed at best. For frame of reference, China is about the same size as the US, but has about 3 times the population, or a total market of ~1.2B people. However, when thinking about our market offerings, therapeutic antibodies, the market shrinks quickly. There is a reality, for better or worse, that of the total Chinese market size, less than half of the population would ever have any chance of affording our products, even at ‘Chinese prices’. Further, of the remaining addressable market, about ½ of that population would likely never get access to emerging therapeutics, even if they could afford them. The reasons for this are varied, but include geography, politics, and limited distribution channels to name a few. So from our perspective, the actual addressable market is roughly the same size as the United States. Still a significant market to be sure, but not a panacea of opportunity.
Expect more on this topic in a week or so…..
Biotechnology Marketing 101 – Part 3
It’s time for more guerrilla marketing for biotech companies. We’ve already talked about social networking in cyberspace as part of an inexpensive marketing strategy, so let’s focus on networking in the Real World.
Networking is a very effective form of Word-of-Mouth marketing. You’re promoting yourself as a business owner or employee and how you do it reflects on the business itself. The best part of chatting socially with other business people is that you aren’t there to close a sale but to cultivate relationships and maybe, eventually, get a few referrals. You don’t need to wear a polo shirt with your company logo (business cards and a name-tag will usually suffice), but you can if the dress code is casual and you don’t arrive decked-out like a NASCAR driver.
How best to proceed? In order to get on the email lists of all the best networking events, you should join a local business association and get involved, especially if there are other science-related companies in town. If you are based near a local University or high-tech business incubator, the opportunities to introduce your company to potential customers is even better. In Madison, Wisconsin, we are lucky enough to have BIOforward as an organization that helps facilitate meeting other science entrepreneurs, potential investors, lawyers and bankers as well as local graduate students who may soon be seeking jobs in our industry. The networking events are a way to meet other biotechnology business professionals in the community. Getting involved in the committees allows you to share your expertise and be recognized by your fellow members. Offer to give a free talk at an upcoming networking event to get the word out on what you do. Everyone needs content, so the event organizers will be happy that you volunteered.
Even if your goal is to build a global brand, your local contacts can help build a buzz about your service or product. These contacts can often be turned into referrals and recommendations for your business, which will help your company grow. In addition, other business owners that you meet might be interested in investing in your company as VCs or Angel Investors. Scientists that you meet at networking events can become customers, collaborators or even employees.
Now, not everyone is a natural at schmoozing, or feels comfortable chatting up their business, no matter how passionate they feel about it. In fact, since a lot of biotech people came out of academic backgrounds they feel better talking about the science rather than the business side. Just remember not to get too technical in your jargon. When a CEOs eyes glaze over, you’ve probably lost them in the minutiae. Listen as much as you speak.
It helps if you know your Product/Service well and have a quick shorthand description of it that you can drop into conversations when people ask the inevitable, “So what does your company do?” Have a 20-30 second ‘elevator speech’ so that anyone can grasp what your business does immediately. Keep everything simple and know how to explain your business in succinct and clear terms. If they want more details about the nuts and bolts of your business, arrange a meeting where you can have their undivided attention.
Relax, chat, listen and bond. And always bring business cards.
Research Works Act
A bill in the US House of Representatives that seeks to prevent federal agencies from requiring open-access publishing of government-funded research would effectively end the NIH’s public access policy. That policy dictates that all NIH-funded investigators submit an electronic version of their final, peer-reviewed manuscripts to the open access resource PubMed Central (PMC) within 12 months after official publication.
The new bill, called the Research Works Act (H.R. 3699), seeks to keep federal agencies from requiring "network dissemination" of research papers without the consent of the publisher.
We at NeoClone oppose the “Research Works Act," as counter to the needs of the scientific community. Biotechnology companies and academic researchers alike require frequent access to published research, new and old. It must be asked; “How would this bill benefit the public?” The current NIH policy is not law, is subject to change, and covers only NIH-funded research. Many, if not all, of the 90,000 manuscripts that are currently made available for free each year will disappear behind a costly subscription wall if this bill is made into law. Legislation that tilts the market in favor of the big publishers is counter-productive. This bill would significantly inhibit scientific discovery and innovation as well as severely limit public access to this information.
We of course believe that publishers have the right to materially benefit from the journals that they produce. However, the one-year grace period is more than enough time to recoup costs of publication, especially since the actual authors of the works in question, the NIH-funded researchers, are not paid for their work by the publishers, but by taxpayers. The peer review process itself is generally performed by uncompensated scientists, and many scientific editors are unpaid as well. With the move to digital distribution of most of the scientific literature, the single greatest cost, that of printing massive quantities of the journals onto paper, is fast becoming a relic of the past.
"Free and open information to scientific literature and data are the underpinnings of discovery in the digital age," Sage Bionetworks' President and Co-Founder Stephen Friend said in a statement put out by the Alliance for Taxpayer Access, a group that advocates for open access to federally funded research results.
Unsurprisingly, representatives of the scientific journals industry have come out in support of the bill.
UPDATE: The sponsors of RWA have withdrawn the bill after an outcry from thousands of researchers. A proposed boycott against publishing group Elsevier, who supported the bill, seems to have helped Elsevier decide to withdraw their support, stating in a letter that its endorsement of the proposal caused some people in the research community to question its commitment to ensuring "the best possible access to research publications and data." More on this: here.
As reported by the Washington Post, House and Senate lawmakers have agreed to reauthorize and enhance two small business research programs that were set to expire this week.
Members of Congress have struck a deal on a defense bill amendment reauthorizing the Small Business Innovation Research program (SBIR) and the Small Business Technology Transfer program (STTR), which require government agencies to set aside a part of their annual research budgets for contracts and grants to small businesses. Both programs are currently running on a temporary resolution that expires Friday. The new amendment would not only reauthorize the program for six years, but also increase the small business allocation requirements for government agencies. Lawmakers also agreed to increase early-stage funding awards and allow small businesses majority-owned by venture capital firms to compete for a small portion of SBIR and SBTT contracts and grants.
NeoClone, like many of its biotech brethren, has received support from these programs over the years. These funds are truly critical to continue to spur innovation in the life science sector. There is no doubt that many interesting and promising projects would never get the necessary momentum without this non-dilutive capital. We are extremely pleased that, even at this time of heightened partisan politics, both sides of the aisle could come together to support this important piece of legislation that will secure funding for innovation and research for the next six years.
The amendment will be attached to the final National Defense Authorization Act, which is expected to be finalized and sent to the White House by week’s end.
Biotechnology Marketing 101 – Part 2
You are a small biotech company. You are pumping every last dollar you’ve squeezed from SBIR grants, angel investors and devilish bankers into overhead, product R&D and production costs, not necessarily in that order. It dawns on you that you don’t really have a marketing budget. What were you thinking, that the world would beat a path to your door starting from Day 1?
Well, if you use guerrilla marketing techniques, they might.
It sounds obvious, but start by having a quality product or service. If you don’t have that, it’ll be hard to generate positive word-of-mouth buzz and impossible to keep customers around. But even if you’re sure you’re offering the next big thing in biotechnology, be it your expertise or superior reagents, how do you get the word out there?
Publish new content to your website and social media frequently, preferably every day but at least once per week. People will be more likely to find your site, and thus your company, due to relevant content and search engines. If they like what they see, they’ll be back. Maybe they’ll tell all their Linked-in and Facebook friends, as well as the biochemistry post-doc down the hall.
What do you post? Well, in addition to highlighting new products and alerting the world to your brilliantly successful round of financing, try repurposing content that you’ve already created to get more marketing visibility. That Powerpoint presentation you gave to potential investors? Post a Reader’s Digest version on your site and mention it on twitter. Digitally record your lab tech demonstrating your top product and post it on that product’s page on your website. And then tout it on Linked-In. Do something unusual, but not too weird. Do you have employees who have a flair for the creative? Produce an inexpensive but entertaining biotech-themed video with jazzy music and maybe it will even go viral. (See some of the top viral science videos here.) Or license a science-related comic strip to bring in the geeks.
Once you have regularly updated content, get the bloggers to write about you, your company and your products or services. Science and Technology bloggers are all over the web. They're always looking for something new and they are a lot easier to contact than reporters from major media outlets. Talk to them about your company, let them know you follow their blog and let them spread the word. And if one of your products gets cited in a journal, no matter how obscure, toot your own horn about it!
Identify your Brand Fans. These are your most valuable and loyal customers. They share positive word-of-mouth about your company, products and services for free, through Facebook, Twitter, Forums and in real life. Do a Google search of your company name to see who is talking about you and what they are saying. Identify the ones who like what you do and be sure to thank them. Ask them for referrals and testimonials that you can put on your website. They’ll love the personal attention.
There are many ways you can market your company on a limited budget. It may not be fast or easy, but it is possible - happy bioteching!
Biotechnology Marketing 101 – Part 1
When you’re a smaller biotech company, you need to use guerrilla marketing techniques to get the word out there about your unique life-science products and services. The 500-lb gorillas in our industry have large advertising budgets and marketing departments, not to mention an army of salespeople constantly storming every lab in the world.
But what about the little guys? Most biotechnology companies out there have 1 to 25 employees and an advertising budget in the zero range. Let’s assume you have a great science-related product or service. But nobody knows about it yet! Word of mouth is one of your most important tools, but it can only get you so far, especially if you’re only using your mouth. Let’s also assume that there are only so many hours in a week and your resources are limited.
So where do you find new life-science customers without having to take out another small business loan?
First, you need a professional-looking website, with a store enabled so that customers can place orders directly. If you are not online, you don’t exist. If you don’t have the ability to put this together in-house with pre-made website templates and dependable web-hosting, then hire somebody to help you. It’s that important. Don’t get too fancy with animation and lots of different colors and fonts. Think about the business websites that YOU find compelling. Add new content regularly and learn (or, again, hire somebody to help you with) SEO, search engine optimization. People are looking for what you sell, so make it easy for them to find you!
While you’re at it, see if you can make a “mobile-friendly” version of your website that can be viewed on smartphones without giving the reader a headache. Companies like SoLoMo can help with that.
Be an Expert. Participate in life science forums such as Scientist Solutions and help out in your area of expertise, whether it is Bioinformatics, Antibody purification, PCR… whatever. Your potential customers are right there, waiting for you to impress them. If you are the business guy and don’t have the technical expertise, then put one of your science hot-shots on this. It doesn’t take a lot of time to post expert advice in these forums. Always remember to have a link back to your website in your online signature. A word of warning: Don’t spam forums with sales pitches! This is an organic method of growing your business and scientists in forums don’t want a hard sell.
Use Social Media. In general, Biotech companies should be tweeting, blogging, and linking-in like crazy. Every other high tech business is doing this, because it works. There are quite a few bioscience-related linked-in groups and twitter feeds that are directly applicable to whatever you do or sell. Smaller biotechs need to squeeze more out of social media and it doesn’t have to take up more than a few minutes per day.
To Be Continued…
Every researcher has them (some more than others). It can be a sudden unexpected discovery or it can be the culmination of months of painstaking work. Most commonly, it's that moment when your long-awaited data comes in and you see the result you've been hoping for; your proof of concept in black and white.
If you are in Academia, you repeat your experiment and rush to publish your results before another researcher half the world away scoops you.
If you are in Industry, you repeat your experiment, then talk to your Director of Research, your Chief Scientific Officer, the CEO and the patent attorney. Along the way, juicy tidbits might make their way to potential and current investors. Or not, depending on the nature of the discovery.
Just last week we had one of those Eureka moments in our research lab, and no, we can't tell you what it was. But the euphoria felt around here was palpable. A difficult problem was solved, and a path forward was established. A point to note was the teamwork behind the effort. Many hands had a part, more so than in many academic labs, where fame is more important than fortune. Because there is no struggle for 1st authorship on a journal article, the goal being a new, valuable product or service, the team effort crossed departments. It helps that everybody knows one another and is working towards the same goal. Innovative research is important in keeping any company that does its own R&D competitive in the marketplace and increasing the value of its products to its customers. Plus, there is an excitement that comes with knowing that your discoveries will soon be used by people in the real world.
That’s why researchers are always happy to be able to exclaim, “Eureka…I have found it!”
Good News for Biotechnology
NeoClone is in support of the National Bioeconomy Blueprint, which is to be released in January of 2012. In the midst of the current economic environment, a strong effort to support biological R&D is welcome news. We’re glad that Washington realizes that “Biological research lays the foundation of a significant portion of our economy.” The Biotechnology industry is ready to expand its highly-skilled and innovative labor force. We produce cutting-edge research and the products that will help provide the answers to many of today’s health and environmental issues. The Blueprint comes at a time when there is a need to reform the regulatory and commercialization hurdles that place huge burdens on scientists, entrepreneurs and investors. Many of the small companies and University start-ups that make up the majority of our industry are at risk of under-capitalization. A large investment in Biological research at the Federal level can help grow the industry, encourage more robust private investment and help these companies thrive. And that’s a result we can all look forward to with anticipation.
Stay tuned for more information on Wisconsin’s own upcoming Biotech-friendly jobs legislation.
Interest grows in therapeutic antibody markets.
Has anyone been paying attention to the amount of financing that novel platforms for generating antibodies (or antibody fragments) that are suitable for therapeutic applications has been getting? The numbers are quite staggering! Here are some examples: Delenex Therapeutics AG, a Swiss biotech company, is developing therapeutic antibody fragments targeting serious medical diseases with high unmet need. These antibody fragments called PENTRA® fragments are being designed for local/topical application by using the benefits of an antibody therapy with only limited systemic exposure. On May 3, 2011, they announced that they raised ~ $35M in Series A funding.
Another example is f-star GmbH, a biopharmaceutical company developing a new generation of antibody based products using its Modular Antibody Technology platform, which recently received ~$21.5M in financing. They have raised ~$49M total so far.
Need more examples? KyMab, with its Kymouse platform technology raised ~$29M within the last year and my personal favorite financing story, Ablexis, who has raised at least $50M from 5 different pharmaceutical companies (in addition to $12M raised from VC land) based on their idea of a better platform to make therapeutic grade antibodies.
That is a total of at least $175M put into antibody platform technologies in the last year! Why is the interest so high? I think two reasons. First, antibodies as drugs are both effective and profitable, making the space in general very investable. Second, there are currently very few companies out there that can do this type of work, largely because they have all been acquired.
Stay tuned as NeoClone positions itself to compete in this very lucrative market in 2011…
When do I know if I have a good antibody?
It is not uncommon in an antibody development project to get exciting results at the bulk culture development stage only to be disappointed later when the clone you were interested in looses expression, based on ELISA data. This happens primarily for one of two reasons. First, there is a chance that the antibody was never really there and the data you are looking at is just artifact antibody – that is antibody that was produced from a cell line that is no longer present. Second, it may be a real antibody, but the expression is so low that it cannot be stably grown out. So that begs the question, what is the chance that the antibody that I am interested in will be successfully sub cloned? In general, NeoClone will have a success rate of 75%+ in subcloning the antibody of interest if the OD in an ELISA is greater than 1. However if the OD is between 1 and 0.5 the success rate drops to ~50%, and if the OD is between 0.25 and 0.5 the chance of a successful subclone drops to about 25%. So while there is always a chance that a clone will not successfully subclone from the bulk culture, NeoClone’s experience would strongly suggest that the higher the OD of the ELISA the better the chance of a successful subclone.
Why do I need more than one good antibody?
Recently I had a discussion with a client that I think is worth sharing. As we discussed project design, I asked what might be the intended end application. They planned to use the developed antibodies in an ELISA as well as in a lateral flow test. So I suggested we increase the number of subclones to be generated because of the multiple end uses....They did not see why this would be the case, "because after all, I just need one good one." However, our experience at NeoClone indicates this is often not the case. Let’s say you find one antibody that works exactly as you want in the ELISA, the assumption is often that the same antibody will be the best candidate for other applications. But, we have numerous examples when an antibody that is great in an ELISA is not so good in a lateral flow application, or vice versa. So what can you do to address this situation? We always recommend taking a number of cell lines through the development process. Unless you know for sure that you will only have one application, the more candidates you have at your disposal, ultimately the lower the cost of application development. It is also worth noting, that if you have designed the project appropriately, to end up with multiple candidates at the end of a project there is very little additional cost than if just one cell line was take to completion. So unless you are positive that you will only need antibodies for the one application, more candidates are always better!
Why is my antibody-producing cell line losing expression?
It is very frustrating to complete an antibody production run and realize that the yield was significantly less than the last time it was done. What happened?
There are a number of possibilities. First, it is possible that the hybridoma has become unstable over time. This can happen if the clone has not undergone enough subclonings. Second, the media conditions may have changed. Hybridoma cell lines will react very differently in different medias. If the initial production run was done in ‘rich’ media, and future runs were done in ‘standard’ media, it would not be at all uncommon to see different yields from the two runs. Also, if the size of production run changed (for example using 4L spinner flasks instead of 1L spinner flasks), the outcome could be very different. Just because you get a certain yield in one system, do not assume that the yield will stay the same in other systems.
So if it is important for you to be able to predict the yield NeoClone would suggest doing an additional subcloning of your clone, documenting the media conditions used and keep the production lot size consistent. If you do this, you should be able to predict yields on a consistent basis.
What yield will my antibody produce?
When we develop antibodies on behalf of clients one of the most common questions we get is ‘what will the yield of the antibody be’? And our answer is always the same – it depends on the specific clone, but if it is a stable producer you should expect between 5 and 50ugs/ml. We understand there is a 10 fold difference in the range of expected outcomes, but our experience suggests this will be the range no matter what you do to the cells in the development stage. In other words, we have not come up with a trick to increase the yields of antibody producing cell lines from the initial subclone. That said, you may see an increase in yield with additional subcloning or by using a richer media, but no guarantees.
So what do you do if a higher yield is an absolute requirement? If this is a priority our suggestion would be to take more clones into production and look for this characteristic. You can do smaller production runs (200ml) with a quick purification that can give you a good idea of what the yield will be with larger production runs. Also, we would strongly recommend developing master banks and working banks of cells to help with consistency and prevent lot to lot variation in yield.
How stable is my clone?
This is always an important issue for monoclonal antibodies – why would you pay for the cost to develop a mAb if you did not think it would be reproducible over time? While the obvious answer is you would not pay if you thought it would be unstable, the reality is that often not all of the work is done to ensure long-term stability. It continues to be our experience that the best way to insure long-term stability is through additional subcloning.
People often cringe at this and ‘hope’ for a different answer. This is frequently because the perceived cost of subcloning is high, but in our experience this is not true. Most often (unless a clone is exhibiting an unstable expression pattern) additional subcloning can be done at relatively low cost and be completed in a couple of weeks. The perception of high costs often comes from a difference of understanding on what it really takes to make great antibodies. If the clone has longer-term importance to you or your company, taking the time (and absorbing the cost) to do the additional cloning is trivial compared to the cost of trying to rescue a clone, or even worse, to replace a lost clone.
So how do you know if your clone is stable? In short, the more the clone has been subcloned, the better the chance that it will be stable over time. This does assume liquid nitrogen storage. Our data generated from tens of thousands of clones would suggest the following regarding clone stability:
If a clone comes from a bulk culture or initial fusion product it is not stable at that point. Even if it exhibits excellent characteristics, there is a strong chance (20%+) that the clone will loose expression after an additional subclone.
If a clone makes it past that subclone, and it is further subcloned, there is a 90%+ chance of long-term stability.
If a clone successfully passes two or more subclonings, there is a 98%+ chance of long-term stability.
So while the answer of clone stability has to come down to your unique situation, our experience would suggest that if the clone has been subcloned two or more times it is likely to be stable for the foreseeable future assuming it has been stored properly.