Author: Alphonse Lab

An immunologist passionate about science and research!

“PREP-TO Qualify” – Tips for Qualification/ Re-class examination

by Alphonse Lab, posted in Education, General, Non-specific, Uncategorized

Prelude

Ma was elated as he looked at the temple from a distance.

As he came close, the edges of the stone stairs welcomed him back into his familiar world.

“These stones are surprisingly uniform”, thought Ma as he took the steps back into the temple.

“Am I learning anything in these journeys?”

Ah! Here is the world I learn a lot from! thought Ma

“What did you learn in this trip?” asked the Zen Master.

Ma looked puzzled and couldn’t hide his frustrations.

“Why do I have to do this?”

“Why do you make me do it every year?”

“You make me travel to my village. I walk the same road, cross the same fields and valleys. I travel the same bridges every year. I anticipate change and gape at even the subtle ones. Without a thought I wave at the children who happily cross by.

Familiarity is breeding boredom, so I am lost in my thoughts! I start to think what would have changed in my village, who would have ceased to dream or even worst whom I know would have ceased to breathe. This is a journey I forebode every year and return back thinking about all this in reverse all over again.”

“What do you expect me to learn? Don’t I learn what I should here in this sacred temple?” blurted out Ma

The journey to Qualify

Do we not learn what we should by doing research? Don’t we all learn in the space between lab and desk? Why should we have to go through this so called “Qualification or reclassification” in order to get a PhD? Didn’t we get admission through selection? Why is that not enough to qualify?

As we prepare for our qualification exam, probably these are some of the question that go through (most of) our minds.

Well, everyone of us know the answer to these questions if we give into the process. I guess, it is within each individual to perceive the potential as we see fit!

Qualification exam is a journey, a passage that we take in the process of getting a PhD. As we prepare to take this exam, the preparation itself becomes a journey and the exam is an experience that aims to mold us as “independent researchers”. “The more we are prepared for this journey the better and richer our experience”.

This article aims to be an aid/tool in this preparation process. It is more general and comes out of my experience. I would urge you to go through this tool and take away things that may resonate with you. Take things that you think may help you. Leave the others. Your journey is unique and if this blog helps, it has met its purpose.

I would like to encompass the complete preparation as

“PREP TO Qualify”

P-R-E-P (Proposal, Review, Examiners, Practice)

DSC_0012_edit_toned_focusProposal:

Proposal is an important document (to describe the project) to the examiners and a lot will depend on its preparation. Nothing new, I know! but it is important to keep in mind as we prepare the document, that this document will give away the plan for the next couple of years and how well you are prepared for the journey.

It is no secret, but a gentle reminder that, “a well defined project will define your qualifying exam”.

  1. Follow instructions:
    Stick to rules, prepare as per required format, do not go overboard.
  1. Preliminary data helps, definitely helps!!
  2. Ask yourself! Does the proposal have the potential to say a story? A story is better received!
  3. Take care in designing the experiments. Know the time it will take to perform and complete the experiments. It is important!
  4. Take the comments from committee members seriously as you prepare the document.

Review:

Review the “area of research” and the subject background. It is what you are tested on! It is important to prepare well by reviewing the material at hand.

  1. Start early to prepare
  2. “Plan your preparation”. Have a strategy to cover extensively the research area and subject background.
  3. Take time to read previous work done in the lab (related and unrelated)
  4. Give enough time for subject background (specialization area)
  5. Cover recent development in the field of research.
  6. Don’t forget to learn about the recent developments in experimental techniques pertaining to the field

Examiners:

  1. Know your examiners.
  2. Learn about their research and techniques in their lab
  3. If they take any courses in the area of your research. Cover the bases
  4. Their focus on research (strengths) and interests.

It is easy to ignore this, but don’t.

Practice:

  1. Give your proposal to peers in the lab. Get their feedback. A good peer will not be judgmental. It is better to “fine tune” your proposal before submitting to the examiners. It never hurts in getting some advice.
  2. Welcome questions from your peers from the lab or outside. Field their questions from the subject and project. It will definitely help on the day of exam.
  3. Practice fielding hypothetical questions, including experimental design ones!
  4. Practice your presentation talk in advance.
  5. Revisit your plan at regular intervals and re-strategize your preparation accordingly.

With PREP under control, it is before the exam and on the day that you have “T-O” (Taking control, On that day) qualify.

Taking Control:

  1. Reduce your stress before your exam
  2. Plan for the day
  3. Practice your talk prior and be comfortable with the slides.
  4. Keep it simple. Remember, the examiners have already read the proposal. So go easy with the presentation.
  5. Remember to relax. Read non subject articles or books.

On the day:

  1. Eat well on the day of exam.
  2. Dress comfortably and know that you are prepared and planned for this day.
  3. Keep a bottle of water during your exam.
  4. Take time (if you need) to answer the questions.
  5. Remember, it is alright to say “I don’t know”. It is better to say “I don’t know” than half guess the answer, because it will avoid further questions from that half guess.
  6. Be prepared to draw figures and be willing to use the chalk board to explain (You may be made to draw on purpose!).
  7. It may be hard, but try to enjoy the process of the exam. After all, you are the expert fielding the questions!

All the best to all who PREP-TO Qualify!

Epilogue:

After hearing patiently Ma’s frustration, the Zen Master turned back to retire to his room.

“Don’t you want to know what I learnt?” asked Ma

“I have!” said the Master

-Author: Martin P. Alphonse

Germ-line: “To edit or not?”

by Alphonse Lab, posted in Specific-Genetics, Specific-Immunotechnology

“Knowledge without wisdom is like an unsaddled horse” – Zen Master

Since the 1970s, the saga of “rDNA technology” has been a steady narrative. In this 40 years of development and evolution, gene editing techniques have become more and more easier and accessible. CRISPR-Cas9 system testifies to this fact. And time after time, every turn of gene editing advancement has witnessed sharp ethical questions fencing its signs.

Rightly so, it should! especially when technologies bare opens the possibility of creating heritable mutations, answers to ethical questions becomes an entitlement.

As I write this post, The US National Academy of Science, Engineering, and Medicine, along with British Royal Society and Chinese Academy of Sciences are co-sponsoring an international summit (in Washington, DC, USA from December 1st -3rd, 2015) to debate and discuss the scientific and societal implications of germ-line genome editing.

Clusterd regularly interspaced pallindromic repeats (CRISPR)-Cas9 system:

About CRISPR-Cas9 technology: 

CRISPR-Cas9 genome editing takes advantage of a prokaryotic acquired immune system. The simplicity of this technology gives the power to alter genomes precisely than ever before. With this system, we could replicate the genetic basis for human diseases in model organisms and scientists can gain insights into the mysterious and puzzling disorders, that was not possible earlier. The technology can be used to correct genetic defects in whole animals as well as in tissues cultured from stem cells. A strategy that has the potential to be eventually used to treat or cure human diseases. The Cas9 enzyme also has the ability to alter epigenetic markers without altering the DNA code, providing a means to manipulate the products of transcription.

Germ-line editing:

DSC_0024_edit_it.71857f089e564c7399626b5977cfb7a7Genome editing in a non productive and fully developed cell have effect only on the treated organism or person (which is not heritable). However, gene editing in germ cells such as that in eggs, sperms or developing embryos can be passed on to the future generations. CRISPR-Cas9 system has the ability to be used in both non productive as well as germ cells (germ-line). And the fact that scientists from Sun Yat-sen University in Guangzhou, China edited human embryos (P. Liang et al., Protein Cell 6, 363-372; 2015) shows that this technology has profound implications for germ-line editing and raises the urgent need to address the social and ethical concerns.

Different voices:

A range of voices, that favor the advancement to outright banning of the technology for foreseeable future can be heard within the scientific community.

 “Encourage the innovators.  says George Church (Geneticist, Harvard Medical School), one of the favoring voices “Banning human germ-line editing could put damper on the best medical research, driving the practice underground” is his argument. Moderate voice such as Jennifer Doudna (Molecular biologist, University of California) suggest “Embryo editing needs scrutiny. and reason that “a complete ban is impractical, given the widespread accessibility and ease of use of CRISPR-Cas9.” Thus, favoring formulation of guidelines for use of the technology.

Scientific research is important in understanding complex genetic diseases. When it comes to creating heritable mutations with editing technologies, it is a fact that we do not know enough about many of the effects (both on and off-target) and its limitations. Complete banning would further limit our understanding. It is here as scientists that we have to tread carefully. Hopefully, the international summit will act as a primer for formulation of guidelines in human germ-line editing.

In agreeing with Jennifer Doudna, formulating guidelines that address

  1. Safety (of the technology),
  2. Communication (with engaging scientific community, ethical bodies and general public),
  3. Guidelines (policy makers and scientists)
  4. Regulation (efficacy and specificity) and
  5. Caution (human germ-line editing for creating genome modified humans should not proceed at this time)

will be key for a progressive way forward.

Reference:

Doudna, J. A. (2015). PERSPECTIVE: Embryo editing needs scrutiny. Nature, 528, S6.

Church, G. (2015). PERSPECTIVE: Encourage the innovators. Nature, 528, S7.

Integrative learning at the lab bench: An approach with Fink’s taxonomy of significant learning

by Alphonse Lab, posted in Education, Non-specific

Prologue

It was supposed to be that moment of Zen. It usually is never a problem for Ma. But today was different!

With his eyes closed and legs crossed, he started to wander in his thoughts.

“Why is this difficult? I am following what I am used to everyday! ”Why is my meditation not fruitful?”

Despondency at the bench:

“The student” is a pivotal cog in the wheel of basic science research. The inevitable pressure of publication and the “result oriented approach” of many principal investigators percolates to their students at the bench. This ever increasing pressure of performance is perhaps one of the many reasons that underpins the increasing despondency for students in basic research. It is here (at the bench) that the students have to balance the ropes of learning and produce publishable results within a limited time. This pressure creates an imbalance for many budding research aspiring students. At the bench, this breach distances the students from learning the skills for a given experiment; Instead, it elbows them to narrowly execute the protocol at hand. Following a protocol without understanding an experiment has far reaching repercussions than mere unproductivity. It tramples on the students learning quotient and converts them into result pursuing engines. So when an experiment fails, the disconnect is further intensified creating an overall negative effect towards basic research. As a result, many students even those with keen aptitude to scientific research get disoriented and lost. Often, it takes valuable time to get back on track.

Can this issue be addressed from the student’s plane? Can students bridge the distance between learning and “result oriented approach” at the bench?

Integrative learning at the lab bench: An approach with Fink’s taxonomy of significant learning is an attempt to answer these questions by taking a leaf from the pedagogical tree. It is an effort seeking to restore a balance at the bench from a student’s learning perspective.

Taxonomy of learning:

Pedagogically, taxonomy of learning serves as a pointer for designing courses. In higher education, depending on the year of study, level of understanding, strength of the students and course’s objectives, taxonomy of learning provides a framework for course designers. One of the well established taxonomy of learning is “Bloom’s taxonomy or Revised Bloom’s taxonomy”. It follows a hierarchical pyramid structure. Starting at the lower end is Knowledge, moving higher up through Comprehension, Application, Analysis, Synthesis and Evaluation.

Instead of a hierarchical system, L. Dee Fink proposed a taxonomy of significant learning that has an integrative approach for learning.

I assume from a student’s perspective; this integrative approach may fit well for learning at the bench.

Significant learning at the lab bench:

Finks
Taxonomy of Significant Learning (Adapted from Fink, 2003)

I reason that protocols are guidelines that depend on human skills and intervention for its effective execution at the bench. The student’s challenge is to develop the skills and intervention techniques. With that outlook, I attempt to impart Fink’s taxonomy in a lab setting, and the following topics are integrative in nature.

Foundational knowledge:

Prior to starting anything at the bench, learning the rationale and the principle of the experiment is not only important but it forms the foundational knowledge. It enhances the understanding of the protocol and further aids in remembering key steps in the experiment.

Application:

Remembering the key steps improves the skill that is required to perform the experiment. As we apply the skill in performing the experiment, thinking critically and adapting practically enhances intervention techniques that is key to the success of the given experiment.

Integration:

Upon completion of the experiment, most of the time we tend to focus on the results. We try to analyze the data and miss some of the important connections that are critical for developing intervention skills. Making connections with the results and the experimental question, its rationale and experimental setup are key aspects of integrative learning at the bench. Importantly, one has to remember that results obtained from one experiment (either positive or negative) is a subset of information that must be connected with its collective sets of data that fits within the larger picture of the project. This integrative learning at the bench is an essential component that aids us in becoming self directed learners. Perhaps this is the most significant learning step at the bench! because as we learn to fit results in the larger picture, we get new direction. If need arises, it helps in designing new experiments to change course or move forward with the project.

Human dimension:

One of the interesting taxonomy of Fink’s significant learning model that differentiates from others, is the human dimension. At bench, the way we perform the experiment provides a window to learn about ourselves and others. For example, our interest in performing the experiment, our commitment to the procedure and most importantly the attitude and perseverance upon failures are few characters of the person that gets personified in this exercise. In bearing a reflective experience and a positive approach to the experiment, we learn about our self and others!

Caring:

Fink adds caring in the taxonomy to include new values and interests that we could develop in the process of learning. At the bench, it could be interests developed in the process of performing an experiment or even after completion that stems from our integrative connections. In this process we develop new interests and take care to learn more, it could be a new technique or a new concept or altogether a new area of study. With interest and care we don’t get intimidated by the task at hand. Essentially, one experiment will lead to another. If we care enough about our experiments, it will never fail to lead us to new paths!

Learning how to learn (LHTL):

LHTL is the ultimate goal for this whole exercise. In this process, the objective is to become self directed learners. At the bench, self directed learning enriches the experience of performing experiments. If we enjoy the experience, it brings us back to the bench with new vigor and enthusiasm either for a failed or a new experiment.  In essence, LHTL trains our perseverance!  that will sustain us in basic science research for long. As we pursue our goals, LHTL will help us to direct and design future experiments within the scope of our project and beyond.

Conscious practice of integrative approach at the bench has the potential to effectively bridge the gap between a student’s learning quotient and productivity. Most importantly, principal investigators should take cognizance and try to create a sustaining environment that encourages and teaches integrative learning at the bench.

Perhaps in the process it will bring down the disconnect and the despondency at the bench. Perhaps students with an aptitude to scientific research will be encouraged to sustain longer in basic research.

Epilogue:

The Zen Master came close to Ma and whispered

“Like life, it is a balance. Find your balance!”

Slowly Ma opened his eyes.

Disturbed yet not disrupted, he closed his eyes again. And after few deep breaths, there it was. The moment he was waiting for. The moment of Zen!

Later that day, Curiosity pushed Ma to the Zen Master

How did you find it? he asked

“Your eyes showed!” said the Zen Master

“But, it is not possible! It was closed, wasn’t it? inquired Ma

“Your eyes were closed, but your anxious eyeballs were not!” said the master.

Reference:

  1. Creating significant learning experiences (Revised and updated) – L. Dee Fink

Epigenetics “train” innate immune memory!

by Alphonse Lab, posted in Specific-Innate Immunity
Prologue

The mound beside the temple graced his seat. And the evening lights of the village from a vantage point set the scene for Zen Master’s daily contemplation.

Ma cautiously approached and sat beside him.

The dim evening lights of the village invited their thoughts and minds.

“How was your day?” asked Zen Master

Trying to recollect his memories,

Ma replied with a smile

“You train me well.”

And after a pause, he continued

“I will remember everything you teach me and one day I will become a Zen Master just like you”

“Can you remember everything?” asked Zen Master realizing the importance of his training

Ma tried to understand the tenor of the question in his mind.

“Can I?” inquired Ma with hesitation, quickly giving up his thoughts.

Immunological memory

Immunology memoryMemory in our immune system is indispensable. Immunological memory is the ability of the immune system to respond more rapidly and efficiently to pathogens that have been previously exposed. Threat from invading pathogens and environmental triggers (henceforth antigens) are constant to the host. And to cope with these threats, immunological memory is pivotal in its recognition and elimination.

Based on the ability to respond to antigenic challenge and contingent on

  1. Specificity and
  2. Clonal selection (theory), which leads to Immunological memory, the immune system is classified into innate and adaptive immunity.

When encountered with an antigenic challenge, adaptive immune cells such as T or B lymphocytes adapt (hence the name adaptive) in response to the antigen. This response is specific and as one of the outcome, memory population (to that specific antigen) are generated. And when re-challenged, the memory response is rapid and robust.

However, innate immune cells such as granulocytes, macrophages, monocytes and Natural Killer (NK) cells were thought to be completely non-specific and lacked memory response.

Identification of pathogen pattern recognition (PPR) receptors in innate immune cells has reasoned the complete non-specificity dogma. It is now appreciated that innate immune system has a semi-specific recognition of pathogen associated molecular patterns (PAMPS) from different classes of microbial pathogens. And now, experimental studies and proposed concepts stack up in support of innate immune memory.

Innate immunologists argue that epigenetics is the key that unlocks the memory of innate immune system!

Epigenetics ”train(ed) innate immunity”:

Epigenetics focuses on cellular and physiological phenotypic changes that are caused by external or environmental factors. In essence, these factors turn on or off the gene expression and affect how the cells read genes instead of being caused by changes in DNA sequence.

When innate immune cells (such as macrophages) are treated with fungal structure β-glucan, positive epigenetic modifications occur in histone regulatory proteins (H3K4me1, H3Kme3 and H3K27ac). These changes are responsible for increased response with gene expression of host defense molecules in innate immune cells when re-stimulated with same or similar stimuli. Innate Immunologists argue that, epigenetic alterations due to exposure of pathogens in innate immune cells in vertebrates can equip them with an “antigen independent memory”. So when re-exposed to same or similar pathogens, there is heightened response. They propose such memory as “trained immunity”.

They insist that epigenetics in innate immune cells leading to memory response makes sense at an evolutionary stand point. Evolutionary conservation theory of “systemic acquired resistance” is proposed in innate immunity. This stems from the fact that innate immune memory is seen as host defense feature in organisms such as plants and invertebrates that lack adaptive immunity (Conventional adaptive immune memory is present only in vertebrates).

Furthermore, role of epigenetic modification in innate immune memory have also been shown in NK cells. Studies have shown that these ‘memory’ NK cells can rapidly degranulate and produce inflammatory cytokines upon reactivation.

It is proposed that these epigenetic modifications of cellular function is at the core of long term training of immunity for innate immune cells.

Other proposed mechanisms that are thought to play a role in innate immune memory include Long-term regulation of Non-Coding Ribo-Nucleic Acids (lncRNA) and metabolic pathways including glycolysis and Warburg metabolism are implicated.

Is memory forever?

Given the specificity and gene rearrangement with central and effector memory cells, adaptive immunity is for lifetime.

How long does the ”trained” innate immune cells have memory? Whether the altered epigenetics is retained after proliferation? Can it be transmitted epigenetically in the germ line (as seen in plants)? are important outstanding questions that remain to be answered.

In summary, classical adaptive immune memory is specific, antigen dependent and is mediated by gene rearrangements. Innate immune cells were thought to be completely non specific which lacked memory response. However, recent studies show that innate immune cells can be trained to acquire memory. Trained innate immune memory is semi-specific, antigen independent and can be mediated through epigenetic reprogramming.

Epilogue

Realizing the confusion in Ma’s voice,

Zen Master filled the silence with his consolations

“Train your mind! Be open to all the environment has to offer. Absorb the harmonious changes and try to preserve the experience in your memory. Your trained mind will adapt as need arises!”

“But, will the environment make me a Zen Master like you?” asked Ma

Waiting for an affirmation from his adoring master, his wanting eyes gazed at Zen Master

His glance spoke thousand words to Ma and he returned his gracious

Smile!

References:

  1. Murphy, K. M. (2012). Janeway’s Immunobiology (pp. 1–892). Garland Science.
  2. Netea, M. G., Latz, E., Mills, K. H. G., & O’Neill, L. A. J. (2015). Innate immune memory: a paradigm shift in understanding host defense. Nature Publishing Group, 16(7), 675–679. http://doi.org/10.1038/ni.3178

Looking at a research article through the lens of the last author!

by Alphonse Lab, posted in Non-specific

Prologue

It was the time of the day to light the candles for prayer. Ma’s trained hands arranged the votive with usual nonchalance. As he was immersed in his mission, he came near the bright red one that he had a soft spot for.

Looking at it, he mused on

“How painstakingly I had scrapped you from bits and pieces in and around this temple. Melted you to shape and here you are! I have crafted and created you.”

As he turned

“Wonderful work” said Zen Master looking over the shoulders of Ma.

He looked at the arrangement of the candles. With a warm smile the Zen Master lit all of them, including the bright red one.

As the light threw away the surrounding darkness, the bright red candle glowed with radiance.

Invited by its beauty, Ma pondered

“Even though I had created, the flame does bring it to life!”

The first and the last:

lens of the last author“Publication is the flame that brings a research work to life!” It is an indispensable ingredient in research. From graduate students to senior research scientists, publication is conventional. When a body of research work is set to publish, the authors take place within the gamut of their contributions. The first author(s) takes away the credit for the work, publication and citations in literature. Rightly so! for the author labors and toils with the research question on bench or at the bedside. Controlled experiments in the lab and or surveys, data collection, analysis and interpretations become the author’s vision and mission for many a year. Often overcoming fear, anxiety and failures; the first author yearns to find joy and enthusiasm until a logical conclusion is established. And in many cases, that logical conclusion is a publication.

Seasoned minds with trained eyes do not fail to recognize an all important author who comes along the list at the end, “The last author”. The content of research, expertise, funding are key contributions the author provides and in many cases even the research question is designed by the last author. For these obvious reasons, the author’s imprint in the publication of the research work is indelible. Indeed! The last author does light the flame that brings the research work to life.

Finding the perspective

We pick up a research article for various reasons (that are relevant to our work or otherwise). General interests, breakthrough research articles, titles or for journal club presentations, the list could be endless. In any given situation, it is important to look through the lens of the last author to get a perspective of the article in hand.

This will serve us two main purpose:

  1. Understanding the complete story behind the research article.
  2. Learning and applying the ideas and concepts to our own work.
Conception and theory (body of research):

Theoretical concepts, experimental approach and contribution towards existing body of literature build on to contrast a seminal work from others. Foundation of ideas and the approach taken by authors to answer a research question becomes vital. To understand the conception of ideas, it becomes critical to study the last author’s previous publications. Many would argue that it is unnecessary, as the article we read provides the necessary background and discuss the implication of the research work. I would have to disagree, because seldom it furnishes the complete picture. When publishing their work the authors are limited by space and style. Therefore, summarizing the complete body of research leading to the conception of ideas are often ignored and more focus is given to explaining the results. Thus, research on the previous publications by the last author in essence will shed light into the rationale and objectives a priori of the work done.

Cohorts and experimental design:

In objective, the peer review process weighs the strength of the data. Furthermore, the last author’s previous publication and collaborations will provide insights to assess the strengths of the experimental design and the cohorts involved (in case of human studies) in the study. This information will aid in the characterization of the study and may provide foresight in understanding the authors subsequent experimental designs.

Funding sources and future directions:

One of the advantages of knowing the funding source is that it can trace us back to the outline of the grant. It can provide information on the projects and concepts the last author is working on and what to expect in the future.

Learning about the content of the research is why we read an scientific article. However, for an unabridged understanding of the article one should find the content’s theory, conception of the idea, experimental design and literature argument. These are key perspectives that could be acquired through the lens of the last author. These perspectives will help us in our own field of research.

Epilogue

It was the next day and the time for prayer was upon them.

After arranging the candles Ma came near the bright red one, with little thought he lit the candle saying

“Let me bring you to life today!”

Yet again the radiance filled the room and Ma was pleased.

As Zen Master entered the prayer hall he saw the candles lit. He smiled at Ma and signaled him to wait.

He disappeared into a small room across the hall and after a few minutes returned with verve.

He came close to the bright red one and covered the candle with a glass chimney and raised the candle to the pedestal above. Ma and Zen Master saw the bright red one’s radiance grow even more and witnessed the new glow filling the entire room.

Both were immersed in the aura of the bright red one. And after a few seconds, collecting his thoughts Zen Master broke the silence and remarked

“It is full of life”

Nano-Immuno-technology

The dawn of Nano-immuno-technology: Applications in Vaccinology

by Alphonse Lab, posted in Specific-Immunotechnology

Prologue

The morning was about to dawn and the radiance of the golden hour filled the sky. Swayed by his overwhelming senses, Ma was standing against the waves of the ocean caressing his feet and its sounds permeating his ears. As he turned around, he saw Zen Master walking towards him

“What did you learn?” he asked

“The massive ocean is so overwhelming, I don’t know where to focus!” replied Ma with concern

Understanding the impatience in Ma’s voice, Zen Master with all his mature wisdom, whispered:

“Search! Slowly try to focus on what interests you. The key is finding your interest. Look around, things you see and the one’s you don’t. Stay with it! Make that interesting point your passion. With time, the passion will become so communicable that you will steadily experience the overwhelming ocean with your mind. And in that oneness, pay attention; the ocean will teach you what you have to learn. Remember, the key is finding your interest!” 

The meeting point of Nantechnology and Immunology

Oceans they are! The discipline of Nanotechnology and Immunology are boundless. Trying to find an interesting point of focus at their confluence is as intriguing as the subject themselves. Therefore, the aim of this short digest is not to cover all the important advancements or possible applications; but rather an attempt to focus at the accent point of the fascinating intersection of both worlds.

Nano-Immuno-technologyA nanometer (nm) is a unit of measurement that equals to one billionth of a meter. The technology that uses the unique properties of objects within the overall size range of 1-1000 nm is called Nanotechnology. Yes, size does matter! Especially when it comes to immune responses. Bio substances that induce an immune reaction, including antigens or pathogen associated molecular patterns (PAMPS) are in nm scale. The physical and chemical properties of a substance such as size, shape, and hydrophobicity determines its ability to induce an immune response. And the ability to engineer properties of substances towards modulating an immune reaction, fields the application of nanotechnology in Immunology. And what an interesting field it is! The last decade or so has seen an exponential advancement in the field of nanotechnology. Limited by scope, this short summary aims to focus on its application in development of vaccines.

Road to Vaccinology via Nanotechnology

Nanoparticles as adjuvants

The goal of an effective vaccine is to enhance the quality and quantity of the adaptive immune response (both the humoral and cellular). To this effect adjuvants like alum are commonplace in vaccines. An adjuvant is a substance that is added to a vaccine to increase the body’s immune response to the vaccine. Nanoparticles in form of nanoemulsions (oil-in-water emulsions) that are composed of solvents and surfactants provide such adjuvant activity. An example in current use is MF59 (Novartis), which consists of squalene oil (a natural 30 carbon organic compound from shark liver oil or amarnath seeds) in combination with polysorbate 80 and sorbitan trioleate. MF59 is licensed as adjuvant in influenza virus vaccines in Europe under the commercial name FLUAD®. MF59 have been shown to be a more potent adjuvant than alum for inducing humoral and Th1 type cell mediated (cellular) immune response. It is interesting to note that MF59 is yet to be approved in the Americas. Apart from nanoemulsions, nanoparticle based vaccine carriers that are in development as adjuvants include biodegradable poly(lactide-co-glycolide) (PLGA) nanoparticles, co-polymer hygdrogels or ‘nanogels’, cholesterol-bearing hydrophobized pullulan (CHP) and cationic lipsosomes. These nanoparticles raise interest in the field due to their size and immunomodulation properties.

Novel vaccine strategies

Nanoparticles as adjuvants may sound unimaginative. Imagine this! How about engineering nanoparticles as complete vaccines? That should sound novel in approach. Right? Well, such vaccines are in development and in preclinical and close to clinical trials. Although some genuine ethical concerns do swell against these vaccines, the immunological outcome looks promising. Virus like Particles (VLPs) and Self Assembling Protein Nanoparticles (SAPNs) are a two such nanoparticles engineered towards development as complete vaccines.

Virus like Particles (VLPs) based vaccines

With their ability to replicate and undergo genetic recombination, viruses are the smallest among pathogens and yet ingenious ones that find ways to evade our defense mechanisms. Taking cues from viruses, Nanotechnologists have developed virus like particles (VLPs) as novel strategies for vaccines. VLPs unique size, shape, uniformity and stability of structures closely resemble viruses. However, they do not contain genetic material and lack replicative abilities. VLPs are nanoparticles (20-100 nm) that can be broadly divided into two categories, Non-synthetic VLPs and Synthetic VLPs.

Non-synthetic VLPs can be designed to contain viral protein subunits that have the ability to form a viral capsid. Currently, there are about 20-30 different enveloped or non-enveloped VLPs in preclinical and clinical development against infectious diseases and cancer. As a strategy, one of the main advantages of VLPs that could be exploited is incorporation of other biological structures such as Toll Like Receptor (TLR) ligands, cell targeting moieties or other biologically active mediators that could enhance the efficiency of vaccines.

Synthetic VLPs are derived by chemical synthesis of lipopeptide monomers that could enhance nanoparticle assembly and stabilize the three-dimensional conformational structure of protein antigens. One promising example of Synthetic VLPs that is currently in development is Lipopeptide-based synthetic VLPs (20-30 nm), that have been used to repetitively display a peptide-mimetic epitope derived from V3 variable loop of gp120 protein of HIV virus.

Self Assembling Protein Nanoparticles (SAPNs)

 Learning from microbial pathogens and VLPs, SAPNs are designed using systemic modeling. By taking advantage of both naturally occurring and synthetically engineered biomolecules, SAPNs are designed to optimize immune responses to vaccine antigens. In essence we could call SAPNs as Version.2 of VLPs. SAPNs can be designed to achieve repetitive antigen display for effective vaccine development. In the process, it can enable conformational presentation of inserted protein epitopes in exposed configuration that could protrude or extend from the surface of the particle after assembly of subunits. About 180 peptide chains can be assembled into a single nanoparticle to form an icosahedral structure. The advantage in this geometry enables better antigen display and cellular activation that could lead to significant increases in the specific production of high-titer, high affinity antibodies directed against the inserted antigen epitopes. A successful example of SAPNs is the incorporation of trimeric coiled-coil epitopes from surface protein of severe acute respiratory syndrome coronavirus (SARS-CoV). These SAPNs has been shown to strongly induce virus-specific neutralizing antibodies in mice after immunization. Similar SAPNs specific to malarial parasite Plasmodium falciparum and P. berghei are in development.

Immunologists consider vaccines as an integral part in prevention and fight aginst infectious diseases. Apart from infectious diseases, advantages of vaccines have found ground in fight against chronic diseases and cancer. At this juncture, application of nanotechnology in form of adjuvants and novel vaccine (VLPs or SAPNs) strategies buoy up the world of vaccinology. Over the long haul, some of the ethical concerns regarding the use of nanoparticles are valid and inflammatory status and long-term effects of these nanoparticles should be extensively studied. Yet, important developments in the field of nanotechnology have already found its application in fields of immunosuppression, transplantation and cancer immunotherapy. Furthermore, exploiting its use in experimental immunology both in vitro and in vivo can provide novel application methods and can potentiate our understanding of our immune system.

Epilogue

 The rays of the morning sun warmed Ma, as he was charged with vim and vigour from Zen Master’s counsel.

Little did he know that he had already stumbled upon his point of interest. I should tell this to the Zen Master he thought, but with a smile returned home thinking to himself

“Tomorrow is another day, yet another morning will dawn to teach more. The ocean is full of lessons!”

References:

(1) Smith DM, Simon JK, Baker JR. Applications of nanotechnology for immunology. Nat Rev Immunol. 2013 Jul 25;13(8):592–605.

(2) Torres Andón F, Alonso MJ. Nanomedicine and cancer immunotherapy – targeting immunosuppressive cells. J Drug Target. Informa Healthcare; 2015 Aug;23(7-8):656–71.

(3) Abuelma’atti MT. Nanotechnolgy: Benefits, risks and ethical issues. Exhibition,    “Innovative Engineering for Sustainable Environment.” IEEE; 2009. 5 p.

Science blogs: "The joy is in the ride!"

Science blogs: “The joy is in the ride!”

by Alphonse Lab, posted in Non-specific

“Why don’t you write science blogs?” quipped Andrey.

I turned the knob of the fountain, reaching for a glass of water. With limited inkling, I started thinking about science blogs. Why should one write science blogs? Could I not spend my time writing a review or focus on that manuscript that is long overdue? Aren’t there other science blogs, what difference does it make? Will it not be one of the millions that are already out there? Is it all worth spending the time and energy in decoding and demystifying scientific jargon? Wouldn’t I complicate more in the process, especially labouring a specialized subject such as Immunology? And so I questioned my case to write blogs. As I closed the knob, a tiny drop of water covered the shiny rim of the tap, holding on to its edges. I witnessed a mundane culmination of “physics in action”. In a split second it had its natural course of fall, in a reaching yet short trip before unifying itself with its own kind to fill my glass of water, in essence the last drop to quench my thirst!

“The joy is in the ride!”

Isn’t thirst for knowledge and ecstasy of discovery the quintessence of doing science? We work our fingers to the bone with experiments, replicate results, and communicate via posters, presentations and publications. Writing is an essential, fundamental and an integral part of scientific research. Yet, it is very little that we do to express our ideas both within and outside the realm of our field of research without perspicacity. When we write manuscripts, reports or grants we are limited by style and format. Occasionally writing exercise in science should be more fun and joyful. Science blogs provide a platform to do exactly that! If our every endeavour aims at achievements and accolades, and perceive time as constraints then there is no joy in doing science and its expression becomes burdensome.

Purists would argue against over simplification and may suggest that misinformation may edge into scientific blogs. Au contraire, in the process of demystifying and decoding jargons we may form solid foundations and contour novel ideas. In due process, specializations become more accessible. Isn’t writing blog the best way to do it? I imagine, it is in this undertaking that most of the learning and exchange of ideas would transpire both within and among peers. Overtime, many blogs may sound redundant but it is certain that they act as a supplement in development of skills, both in content and communication. I would tie up that interest in science would prevail at the end of the day.

Finally, a drop of water may not be able to separate from others once it is unified, but the joy for the drop must be in its journey. The journey starting from the tip of the fountain until it joins the many others at the rim of the glass it contains! I reckon, so would be the exercise of scientific blogging “the joy is in the ride!”

Thanks Andrey, “I have started my journey!”