In this blog post, we delve into the distinct advantages of using fluorescent liposomes in cellular studies. Fluorescent liposomes, essentially lipid vesicles embedded with fluorescent probes, have emerged as an invaluable tool in biomedical research. They offer unparalleled advantages in terms of real-time tracking, targeted delivery, and visualization of cellular processes at a microscopic level.

From providing unique insights into the intricate workings of cells to enabling precise drug delivery, these luminescent nanostructures hold great promise.

Introduction Fluorescent Liposomes

Fluorescent liposomes represent a significant advancement in cellular studies, providing researchers with an effective tool for visualizing and tracking biological processes. Liposomes are self-assembling spherical vesicles composed of phospholipid bilayers, which can encapsulate various substances, including drugs, proteins, and nucleic acids. They mimic the natural structure of cell membranes, making them ideal carriers for targeted drug delivery.

Fluorescent liposomes differ from regular liposomes as they contain fluorescent markers or probes within their structure. These fluorescent molecules emit light upon excitation, allowing researchers to visualize the liposomes under a fluorescence microscope. This ability to "light up" under specific conditions provides a crucial advantage in cellular studies.

Fluorescent liposomes enable real-time tracking of the liposomes' movement within cells or across biological barriers, providing valuable insights into cellular interactions, drug delivery mechanisms, and disease progression. These unique properties make fluorescent liposomes a powerful tool in biomedical research, with potential applications in drug development, diagnostics, and therapeutic strategies.

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Fig 1: Liposome–cell interaction.

The Role of Fluorescent Liposomes in Cellular Studies

In cellular studies, fluorescent liposomes play a pivotal role due to their unique properties and versatile applications. They serve as excellent tools for visualizing and understanding the complex processes within cells. Here are some key roles they play:

• Drug Delivery Studies: Fluorescent liposomes enable researchers to track the delivery of drugs encapsulated within these vesicles. By monitoring their movement and interaction with target cells, scientists can optimize drug delivery strategies.
• Cellular Interaction and Uptake: The fluorescent markers within liposomes allow real-time observation of how cells interact with these carriers. This is crucial in understanding cellular uptake mechanisms.
• Biological Barrier Crossing: Fluorescent liposomes can be used to study how substances cross biological barriers, such as the blood-brain barrier, which is essential in developing treatments for diseases like Alzheimer's and Parkinson's.
• Disease Progression and Treatment Response: By tagging liposomes with specific fluorescent markers, researchers can monitor disease progression and assess the effectiveness of treatments.

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Fig 2: Schematic representation of the different types of liposomal drug delivery systems.

Fluorescent liposomes, with their luminescent properties and biocompatibility, have thus become indispensable tools in cellular studies, offering invaluable insights into cellular processes and drug delivery mechanisms.

Advantages of Using Fluorescent Liposomes

Fluorescent liposomes offer several advantages in cellular studies, revolutionizing the way researchers investigate biological processes and develop therapeutics. Here are some of the key benefits:

• Real-Time Tracking: Fluorescent liposomes can be monitored in real-time as they interact with cells or cross biological barriers, providing immediate feedback on their behavior.
• Improved Visualization: The fluorescent markers within liposomes enhance visualization under a microscope, allowing for precise tracking and imaging of cellular processes.
• Targeted Drug Delivery: Liposomes mimic cell membranes, improving their ability to deliver drugs directly to targeted cells, reducing side effects and increasing drug efficacy.
• Versatility: Fluorescent liposomes can encapsulate both hydrophilic and hydrophobic substances, making them versatile carriers for various types of drugs.
• Biocompatibility and Reduced Toxicity: Liposomes are composed of biocompatible materials, reducing the risk of toxic reactions that may occur with other delivery systems.

The use of fluorescent liposomes thus presents immense opportunities for advancing our understanding of cellular behavior and improving therapeutic strategies.

Fluorescent Liposomes vs. Traditional Methods

Fluorescent liposomes present a considerable leap forward when compared to traditional methods in cellular studies, particularly in drug delivery and visualization of cellular processes.

Traditional methods often involve the direct administration of drugs or the use of non-specific carriers that lack targeted delivery. This can lead to suboptimal therapeutic effects and increased side effects. Fluorescent liposomes, on the other hand, can encapsulate and deliver drugs directly to the target cells, improving therapeutic efficacy while minimizing side effects.

In terms of visualization, traditional staining and imaging techniques can be invasive and may not provide real-time tracking of cellular processes. Conversely, fluorescent liposomes enable real-time observation and precise tracking of cellular interactions and biological barriers crossing due to their inherent luminescent properties.

Moreover, traditional methods may not provide the versatility that fluorescent liposomes offer. Liposomes can encapsulate both hydrophilic and hydrophobic substances, broadening the range of potential therapeutics.

In summary, fluorescent liposomes outperform traditional methods in several aspects, making them a superior tool in cellular studies and drug delivery research.

Future Prospects of Fluorescent Liposomes

Fluorescent liposomes hold immense potential in the future of biomedical research, promising to revolutionize our understanding of cellular processes and the development of novel therapeutics. However, it's crucial to note that products such as those offered by FormuMax are intended solely for research purposes and are not designed for clinical or therapeutic use.

Targeted Drug Delivery: One of the most promising prospects of fluorescent liposomes lies in their potential for targeted drug delivery. By improving the specificity and efficacy of drug delivery, fluorescent liposomes could pave the way for more effective treatments with fewer side effects.

Improved Diagnostics: Fluorescent liposomes could also significantly improve diagnostic techniques. Their ability to detect changes in pH, temperature, and the presence of specific biomolecules offers vast potential for the development of advanced biosensors and diagnostic tools.

Personalized Medicine: The application of fluorescent liposomes in personalized medicine is another exciting prospect. They could be used to deliver personalized drug combinations, tailored to the patient's unique genetic profile and disease state.

Neurological Research: The ability of fluorescent liposomes to cross the blood-brain barrier opens new avenues for neurological research. This could lead to breakthroughs in the treatment of brain diseases, such as Alzheimer's and Parkinson's.

Vaccine Development: In the field of immunology, fluorescent liposomes have shown potential as vaccine adjuvants, enhancing immune responses and improving vaccine efficacy.

In conclusion, while fluorescent liposomes have already revolutionized biomedical research, their future prospects are even more exciting. As we continue to explore their potential, fluorescent liposomes will undoubtedly continue to shape the future of medicine.

This blog post aims to dissect the science behind EMLs, exploring their distinctive characteristics, their advantages over conventional liposomes, and why they are poised to revolutionize drug delivery. We will delve into the biology that underpins these nano-scale vesicles, shedding light on their ability to potentially overcome some of the most significant challenges in modern medicine. Buckle up for an enlightening journey into the world of advanced drug delivery systems.

Understanding Exosomes

Exosomes are naturally occurring extracellular vesicles, typically 30-150 nanometers in size, secreted by almost all types of cells. They play a crucial role in cell-to-cell communication by transferring proteins, lipids, and nucleic acids from one cell to another, thereby influencing the recipient cell's function.

Their unique ability to cross biological barriers, including the blood-brain barrier, and their inherent biocompatibility and low immunogenicity have sparked interest in their potential as natural drug delivery vehicles. However, the complexity involved in isolating exosomes and modifying them for therapeutic use has been a significant roadblock.

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Fig 1: Cellular journey of internalized exosomes and endogenously produced exosomes.

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Fig 2: Biogenesis and identification of exosomes.

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Fig 3: Cellular uptake of therapeutic exosomes.

Liposomes, on the other hand, are artificially prepared vesicles made of lipid bilayers. They have been widely used in drug delivery due to their ability to encapsulate both hydrophobic and hydrophilic drugs and deliver them to target sites. However, conventional liposomes lack the specificity and efficiency of exosomes, leading to the development of Exosome-Mimicking Liposomes (EMLs).

Read more about Liposomes: Liposomes: Their Structure and Biomedical Applications

Exosome-Mimicking Liposomes (Mikosome™) Explained

Exosome-mimicking liposomes (EMLs) represent an innovative approach in the field of drug delivery systems. They are artificially synthesized vesicles, engineered to mimic the natural properties of exosomes while overcoming their limitations.

The process of creating EMLs involves two primary steps: First, liposomes are prepared through techniques like thin-film hydration or reverse-phase evaporation. Then, these liposomes undergo a series of extrusion steps through filters with decreasing pore sizes. This process results in liposomes that not only resemble exosomes in size but also in composition and function.

Here are some key features of EMLs:

1. Size and Shape: EMLs are typically 30-150 nanometers in diameter, closely mimicking the size of natural exosomes.
2. Composition: They possess a lipid bilayer structure similar to exosomes, allowing them to carry both hydrophilic and hydrophobic substances.
3. Function: Like exosomes, EMLs can cross biological barriers and deliver their cargo directly to target cells.

In essence, EMLs combine the best of both worlds – the specificity and biocompatibility of exosomes, with the ease of production and modification associated with liposomes.

Mikosome™ vs. Conventional Liposomes

The comparison between Exosome-Mimicking Liposomes (EMLs) and conventional liposomes is vital to understanding their respective roles in drug delivery systems.

Conventional Liposomes have been widely used for drug delivery due to their biocompatibility, ability to encapsulate a diverse range of drugs, and potential for modification. However, they often face challenges such as rapid clearance by the immune system, instability in the bloodstream, and lack of targeted delivery, which can lead to off-target effects and diminished therapeutic efficiency.

On the other hand, EMLs are designed to overcome these limitations. Here are some key advantages of EMLs over conventional liposomes:

1. Enhanced Stability: EMLs are more stable in the bloodstream, which extends their circulation time and increases the chance of reaching the target cells.
2. Targeted Delivery: Similar to natural exosomes, EMLs can deliver their cargo directly to specific cells, reducing off-target effects.
3. Reduced Immunogenicity: As EMLs mimic the properties of exosomes, they are less likely to be recognized and cleared by the immune system.

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Fig 4: Comparison of exosome-mimicking liposomes with conventional liposomes for intracellular delivery of siRNA

In conclusion, while conventional liposomes have their merits, EMLs offer a more precise, efficient, and safer alternative for drug delivery.

Mikosome™ in Drug Delivery

Exosome-mimicking liposomes (EMLs) have shown significant promise in the domain of drug delivery. By leveraging the unique advantages of both natural exosomes and artificially synthesized liposomes, EMLs are paving the way for enhanced drug delivery systems.

Several notable features make EMLs an excellent vehicle for drug delivery. They exhibit superior stability in the bloodstream, targeted delivery capabilities, and reduced immunogenicity compared to conventional liposomes. Their ability to cross biological barriers, such as the blood-brain barrier, enables them to deliver therapeutic agents to specific cells or tissues.

EMLs can be employed to carry a variety of substances, including small molecules, proteins, peptides, and even nucleic acids for gene therapy applications. Additionally, the encapsulation of drugs within EMLs can protect them from degradation, thereby prolonging their half-life and enhancing their bioavailability.

It's important to clarify that the Mikosome™ provided by FormuMax are intended strictly for research purposes. They are not approved for clinical or therapeutic use. They serve as a valuable tool for scientists investigating targeted drug delivery and related areas of therapeutic research.

While EMLs hold substantial potential, more research is required to fully understand their capabilities and limitations in a clinical context.

This innovative marriage of materials science and biomedicine has opened new avenues for treating a range of complex disorders, from autoimmune diseases to various forms of cancer. In this article, we delve into the inner workings of Clodronate Liposomes, exploring their creation, action mechanism, and application in healthcare.

Understanding Clodronate Liposomes

Clodronate liposomes, a groundbreaking development in medical science, have shown immense potential in targeted drug delivery for treating various diseases. The foundation of their function lies in the unique properties of Clodronate, a bisphosphonate known to inhibit bone resorption, and liposomes, lipid-based vesicles that act as drug carriers.

The encapsulation of Clodronate within liposomes facilitates a precise delivery system. Upon ingestion by macrophages, a type of white blood cell, the liposomes release Clodronate into the cells. Here, Clodronate is metabolized into a non-hydrolyzable ATP analog, which inhibits the mitochondrial ADP/ATP translocase—an enzyme responsible for ATP and ADP exchange across the mitochondrial membrane. This process depletes cellular ATP, triggering apoptosis, or cell death.

The specificity of this mechanism allows for the selective destruction of certain cells like osteoclasts and macrophages, minimizing damage to healthy tissues. This has been particularly beneficial in conditions where these cells contribute to disease progression, such as autoimmune diseases and certain cancers.

For instance, in autoimmune diseases like multiple sclerosis, clodronate-loaded liposomes have demonstrated promising results by effectively managing the patient's immune response and reducing the severity of the disease. They also show potential in other medical fields. Cardiac regeneration studies have linked macrophage depletion through clodronate liposomes to compromised cardiac repair. In liver transplantation, they've been found to influence Kupffer cell polarization.

These insights highlight the intriguing nature of clodronate liposomes and their potential for therapeutic interventions. As research continues to uncover more about their workings, we may discover even broader applications for this innovative treatment modality.

The Promising Role of Clodronate Liposomes in Clinical Applications and Autoimmune Diseases

Clodronate liposomes, with their unique ability to target and deplete macrophages, have opened up a multitude of applications in both the clinical setting and the management of autoimmune diseases. These innovative therapeutic tools are especially effective where macrophages contribute to disease progression, such as autoimmune diseases and certain types of cancer.

In the realm of autoimmune diseases, clodronate liposomes have shown immense potential. For instance, in Rheumatoid Arthritis (RA), an inflammatory disease marked by joint inflammation and destruction, they target synovial macrophages. These cells, known to release pro-inflammatory cytokines and promote osteoclastogenesis, contribute to bone erosion. Studies have indicated that intra-articular injections of clodronate liposomes can lead to a significant reduction in synovial inflammation and improved joint function.

Similarly, in Multiple Sclerosis (MS), a disease affecting the central nervous system, clodronate liposomes have shown promise in mitigating the aggressive immune response by selectively depleting harmful macrophages. This results in reduced inflammation and slowed disease progression. Additionally, research suggests that clodronate liposomes could also play a role in managing Systemic Lupus Erythematosus (SLE) by decreasing macrophage infiltration in the kidney.

Beyond autoimmune diseases, clodronate liposomes have also demonstrated potential in oncology. In metastatic breast cancer, they've been used to prevent bone metastases by inhibiting osteoclasts, a type of bone cell that facilitates metastasis when co-opted by cancer cells.

Furthermore, investigations are ongoing into the use of clodronate liposomes in reducing ischemia-reperfusion injury in organ transplantation. By depleting Kupffer cells in the liver, these liposomes could potentially mitigate the inflammatory response and improve transplantation outcomes.

As our understanding of diseases evolves, so too will the applications of clodronate liposomes. This paradigm shift in treatment offers a more targeted and potentially safer option, holding great promise for improving patient outcomes across a range of clinical scenarios.

Clophosome®: A Market Leader in Clodronate Liposomes

FormuMax is a leading name in clodronate liposomes with its innovative products: Clophosome® and Clophosome®-A. These products have been designed to leverage the therapeutic potential of clodronate liposomes, offering an effective solution for conditions where macrophage depletion is beneficial.

Clophosome®, encapsulated with neutral liposomes, is capable of depleting 80-90% of macrophages in the spleen after a single intravenous or intraperitoneal administration. This underscores its efficacy as a potent macrophage-depleting agent.

Clophosome®-A, the second-generation product, is made up of anionic lipids. It surpasses its predecessor by depleting over 90% of spleen macrophages after a single intravenous injection. This improvement in efficacy can potentially enhance treatment outcomes in diseases like rheumatoid arthritis, multiple sclerosis, and certain cancers.

Both products are manufactured under an endotoxin-controlled process, ensuring safety and superior activity. Additionally, they offer a longer shelf-life and ease of use.

FormuMax offers these clodronate liposomes in various combo kits and vial sizes (2 mL and 10 mL), catering to different needs. For applications requiring high doses of clodronate in small volumes, such as intratracheal, intranasal, Intracerebroventricular (ICV), or intratumoral injections, high-potency Clophosome® (20mg/mL) is also available through FormuMax.

FormuMax also offers lyophilized Clophosome® (both neutral and anionic forms) with an exceptional shelf life. It can be easily reconstituted to regenerate the original Clophosome® product, providing a practical solution for long-term storage and usage.

In this blog post, we aim to shed light on the fascinating structure of liposomes and discuss their diverse biomedical applications. With a structure akin to the cell membrane itself, liposomes have opened new avenues in drug delivery systems and more. Stand by as we navigate through the exciting landscape of liposomal science.

Understanding Liposomes: An Overview

Liposomes are spherical vesicles characterized by one or more concentric lipid bilayers enclosing discrete aqueous spaces. They were first discovered in the early 1960s by British hematologist Dr. Alec D. Bangham, who noticed that phospholipids formed a spherical shape when suspended in water.

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In essence, liposomes mimic the structure of a cell membrane:

• Phospholipid Structure: Phospholipids, the primary component of liposomes, have a 'head' that is attracted to water (hydrophilic) and two 'tails' that repel water (hydrophobic). When placed in an aqueous solution, these molecules spontaneously arrange themselves into a bilayer with the heads facing the water and the tails hidden in the center, forming a barrier between the inside and outside of the vesicle.
• Bilayer Formation: The self-assembly of these phospholipids results in a bilayer structure. This bilayer can encapsulate both water-soluble and fat-soluble substances, making liposomes excellent carriers for various types of drugs.

The size of liposomes can vary from very small (nanometer range) to large (micrometer range), which affects their drug encapsulation efficiency, circulation time in the body, and the route of administration.

Understanding the intricate structure of liposomes has paved the way for their use in diverse biomedical applications. In the following sections, we will delve into some of these applications and explore how they are revolutionizing the field of medicine and research.

The Unique Structure of Liposomes

Liposomes are microscopic vesicles that boast a unique structural complexity, which contributes to their functional versatility in the field of biomedical applications. The structure of liposomes is primarily composed of lipid bilayers, similar to the structure of a cell membrane.

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Lipid Bilayer: The Fundamental Unit

The lipid bilayer is the fundamental unit of a liposome. This bilayer is formed from phospholipids, which are amphipathic molecules possessing both hydrophilic (water-attracting) and hydrophobic (water-repelling) properties.

• The hydrophilic or "head" region of the phospholipid is attracted to water.
• The hydrophobic or "tail" region, composed of fatty acid chains, repels water.

When these phospholipids are introduced into an aqueous environment, they spontaneously arrange themselves into a bilayer formation, with the hydrophilic heads facing the water and the hydrophobic tails hidden in the center. This self-assembly process results in the formation of a closed spherical vesicle - the liposome.

The Vesicle: Single vs. Multiple Bilayers

Depending on the number of lipid bilayers, liposomes can be categorized into unilamellar and multilamellar liposomes:

• Unilamellar liposomes have a single lipid bilayer and can further be classified into small unilamellar vesicles (SUVs, less than 100 nm in diameter) and large unilamellar vesicles (LUVs, greater than 100 nm in diameter).
• Multilamellar liposomes consist of multiple concentric lipid bilayers and are often larger in size.

Significance of the Liposome Structure

The unique structure of liposomes offers two distinct compartments for encapsulating substances: one in the aqueous core and the other within the lipid bilayer itself. This allows liposomes to carry both hydrophilic and hydrophobic substances, making them ideal candidates for drug delivery systems.

Indeed, the structural intricacy of liposomes not only makes them fascinating from a biological standpoint but also presents a host of potential applications in the realm of biomedicine.

Biomedical Applications of Liposomes

Liposomes, due to their unique structure and biocompatibility, have been widely used in the biomedical field to enhance the efficacy and safety of therapeutic agents. Here are some compelling ways in which liposomes are currently being utilized in medicine and research:

Drug Delivery Systems: Liposomes have emerged as a promising tool for drug delivery. Their ability to encapsulate both hydrophilic and hydrophobic drugs, shield them from degradation, and target specific cells or tissues, has revolutionized therapeutics. For instance, liposomal doxorubicin (Doxil), an FDA-approved chemotherapy drug, has significantly reduced cardiotoxicity compared to conventional doxorubicin.

Immunology: Liposomes can act as potent adjuvants, enhancing the immune response to antigens. They can carry both antigens and immunostimulatory molecules, thereby modulating the immune response in a desirable manner.

Diagnostic Imaging: Liposomes loaded with contrast agents have been used in diagnostic imaging. The encapsulation improves the stability of the agent, prolongs the imaging time, and can be targeted to specific tissues.

These applications highlight the versatility of liposomes in the biomedical field. As our understanding of liposomes continues to deepen, we can expect more innovative applications that will further improve patient care and outcomes.

Challenges and Opportunities in Liposome Technology

Liposome technology, although promising, presents distinct challenges and opportunities. A significant hurdle is the efficient translation of targeted liposome technology into practical applications. Factors such as liposome-protein corona in physiological environments are challenging to negotiate, yet they hold potential for targeted nanomedicine delivery.

Commercial pharmaceutical liposome applications also pose their unique challenges, requiring a basic familiarity with liposome technology and terminology. Despite these obstacles, liposome technologies have shown potential for a broad range of active substances for injection.

The delivery of phytochemicals by liposome cargoes has seen recent progress, yet it faces challenges such as liposomal stability and permeation. However, advances in technologies for preparing liposomes, particularly ligand-functionalized liposome formulations, are pushing us closer to clinical translation in targeted cancer therapy.

Despite considerable technological success in cancer nanomedicine, there remains a need for further progress in improving PK and biodistribution. These challenges, however, are not without their silver lining. They present opportunities for scientific exploration and advancement in the field of liposome technology.

Seizing these exciting opportunities requires brilliant minds, groundbreaking experiments, and dependable partners like FormuMax Scientific. For the past 15 years, FormuMax has been a dedicated provider of superior-quality liposomal reagents, continually aiding in the advancement of scientific discovery.

Today (03/16/2020), 6 counties in the Bay Area of California, including Sunnyvale where FormuMax is located, announced a Public Health Order requiring 6.7 million people in our area to stay home except for essential needs. This unprecedented step is being taken to try to contain and slow the spread of the coronavirus which causes the COVID-19 disease.

This Public Health Order, which takes effect at midnight, March 17^th, 2020, and is currently scheduled to last until April 7^th, 2020, means that FormuMax is obligated to close our business. As a result, we will be unable to fulfill or ship any new or on-going orders while the mandatory shutdown is in place.

While the weeks ahead have some uncertainty, we pledge to stay in close contact with you as we get more information about when this shutdown will be lifted and normal shipping operations can resume.

We apologize for any inconvenience during these extraordinary times, and thank you for your understanding. Most of all, we wish you all to stay safe, and together, we are confident the world will overcome this significant challenge.

I am very sorry for any inconvenience that may be causing. Don’t hesitate to let us know if we can provide any more information or assistance.

Kind regards,

Peter Zhang, Ph.D.

President & CEO

FormuMax Scientific, Inc.

Publications on Clophosome® (neutral)

Publications on Clophosome® - A (anionic)

Publications using Doxoves® (liposomal doxorubicin HCl)

Publications on fluorescent liposomes

Publications using FormuMax's plain liposomes

The rods inside the liposomes are the doxorubicin/sulfate complexes in a semi-crystal form. 

Doxoves™ exhibits comparable physical characteristics regarding particle size (approx. 85nm), narrow distribution, and drug encapsulation efficiency (>98%), and uses the same bulk buffer solution (10wt% sucrose, 10mM histidine, pH 6.5). Doxoves™ is sterile filtered and filled in autoclaved glass vials for long-term stability. However, Doxoves™ is provided at 4mg/mL drug concentration (compared to 2mg/mL Doxil®) in order to allow researchers to conduct experiments at much higher drug concentrations/doses.

Below you will find some results which we have obtained from research with Doxoves™. Cryo-TEM conducted at NanoImaging Services showed consistent particle size, predominantly unilamellar structure and the rod-like structures of the doxorubicin/sulfate co-crystals inside the liposomes. Results from a recent PK study in rats showed a similar plasma drug PK profile to that of Doxil® with a plasma drug half-life of 30 - 40 hrs. The two batches of Doxoves™ showed identical PK profiles and parameters. You will also find a typical certificate of analysis of Doxoves™ here.

However, please keep in mind that although we have confidence in the quality of Doxoves™, it nevertheless is a research grade liposomal doxorubicin product. It is not manufactured under cGMP conditions. We have not conducted any investigations regarding its biodistributions in tissues and/or in tumors, toxicity and/or antitumor efficacy in house or sponsored by FormuMax. 

Pharmacokinetic studies of Doxoves™ in Rats

Cryo-TEM of  Doxoves™ (Cryo-TEM conducted by Nanoimaging Services)                             Cryo-TEM of Doxil (link to origin)