Cell Membrane-Coated Nanoparticles for Precision Medicine: A Comprehensive Review of Coating Techniques for Tissue-Specific Therapeutics

Nanoencapsulation has become a recent advancement in drug delivery, enhancing stability, bioavailability, and enabling controlled, targeted substance delivery to specific cells or tissues. However, traditional nanoparticle delivery faces challenges such as a short circulation time and immune recognition. To tackle these issues, cell membrane-coated nanoparticles have been suggested as a practical alternative. The production process involves three main stages: cell lysis and membrane fragmentation, membrane isolation, and nanoparticle coating. Cell membranes are typically fragmented using hypotonic lysis with homogenization or sonication. Subsequent membrane fragments are isolated through multiple centrifugation steps. Coating nanoparticles can be achieved through extrusion, sonication, or a combination of both methods. Notably, this analysis reveals the absence of a universally applicable method for nanoparticle coating, as the three stages differ significantly in their procedures. This review explores current developments and approaches to cell membrane-coated nanoparticles, highlighting their potential as an effective alternative for targeted drug delivery and various therapeutic applications.


Introduction
Nanoencapsulation for in vivo administration provides numerous benefits, such as enhancing effectiveness and safety by protecting the substances from degradation or elimination [1,2].This technique contributes to increased absorption and improved bioavailability, optimizing distribution, and extending circulation time while simultaneously reducing toxicity [2,3].Some nanomaterials offer advantages such as enhanced solubility and loading capacity, improved delivery efficiency, and protection from degradation due to the stability provided by the nanocarriers [1][2][3].However, nanoparticle delivery has many limitations.Nanocarriers are very prone to interact with biomolecules in the bloodstream, creating the so-called "biocorona" [4], which results in recognition by the immune system [5].Upon arrival to the target cells, many nanocarriers are trapped in endocytic vesicles and end up being degraded by lysosomes, diminishing the drug delivery efficiency [6].
Recent studies suggest that nanocarriers show an average efficiency of delivering to the desired target of less than 1% [7,8], leaving space for a significant improvement in targeted delivery.As a result, nanoparticles coated with cell membranes have been proposed as a way to address these problems, as they show a combination of the advantages present in natural nanomaterials such as cell membrane-derived nanomaterials, and artificial nanocarriers, such as the aforementioned polymeric or inorganic nanocarriers [9][10][11][12].
Cell membrane-coated nanoparticles are biomimetic nanoparticles that are constituted by a cell membrane cover and synthetic nanoparticles [5].They offer several advantages over bare nanomaterials, such as increased biocompatibility, due to the similarity of biological membranes to cellular materials, reducing the risk of immune system rejection [13].The presence of biological membranes enhances biodistribution by guiding nano-vectored materials to target cells, utilizing membrane receptors recognizable by the target cells.This aspect represents a significant area of study, applicable to immune system cells [13], central nervous system [14], as well as a large number of cancer cells (Table 1).Additionally, coated nanocarriers demonstrate improved drug release control and efficiency, as the biological membranes can degrade or fuse with target cells, releasing the drug at the desired location [9,15].Specifically, the biological camouflage provided by these membranes protects nanoparticles from the body's defense systems, extending their lifespan and reducing the risk of premature elimination [13,[15][16][17][18][19][20].The ability to target particles to specific cells, facilitated by the presence of receptors on biological membranes, is a key advantage that positions nanomaterials coated with biological membranes as a promising option for targeted delivery.for obtaining cell membrane fragments.We provide detailed insights into the processes involved in isolating these membranes and coating nanoparticles with them.The ultimate goal of this review is to examine technology which has been developed recently to generate cell membrane-coated nanoparticles, showcasing their potential for achieving tissue-specific targeting.This review aims to clearly outline the significance of the study within the broader context of this emerging field.

General Procedure
To obtain cell membrane-coated nanoparticles three pivotal and indispensable steps must be undertaken.These steps encompass the cell lysis and fragmentation of the membranes, the isolation of these membrane fragments, and the coating of the selected nanocarriers (Figure 1).

General Procedure
To obtain cell membrane-coated nanoparticles three pivotal and indispensable steps must be undertaken.These steps encompass the cell lysis and fragmentation of the membranes, the isolation of these membrane fragments, and the coating of the selected nanocarriers (Figure 1).
The choice of materials for each of these crucial steps depends on the specific tissue being targeted and the nature of the treatment under investigation.The selection is tailored to optimize compatibility with the intended biological environment and enhance the efficacy of the experimental approach.

Membrane Donor Cells
The selection of a specific cell type is contingent upon the target tissue or application.Typically, cancer cells are employed to specifically target the corresponding cancerous tissue, while white or red blood cells may be used for applications with less specific targets.Most of these cell types were employed to facilitate the precise targeting of nanoparticles to specific tissues.However, some of these cells served a dual purpose by inducing immune stimulation against cancer.
Cervical and ovarian cancer cells were used to favor the cytosolic delivery of cargo inside living cells [21] or for homologous targeting [22].Multiple myeloma cells were chosen to target their equivalent counterparts, ensuring specificity in cargo delivery [25].In the case of melanoma cells, their use was geared towards promoting the delivery and internalization of therapeutic or antigenic materials [12], or for photoimmunotherapy [26].Leukemia cells were employed to deliver cargo into leukemia cells [34] or were genetically modified to express a protein that can specifically target a tissue [36].Neuroblastoma cells were employed for their capacity to capture neurotoxins effectively [79].Breast cancer cells were used to target homologous cells and deliver cargo [6].Similarly, colon carcinoma [57] head and neck squamous cell carcinoma [58], lung cancer [54], glioma [63,64], glioblastoma [65,66], prostate cancer [67], and liver cancer [68] cells were selected for homologous targeting, ensuring precision in cargo delivery to specific tissues.The choice of materials for each of these crucial steps depends on the specific tissue being targeted and the nature of the treatment under investigation.The selection is tailored to optimize compatibility with the intended biological environment and enhance the efficacy of the experimental approach.

Membrane Donor Cells
The selection of a specific cell type is contingent upon the target tissue or application.Typically, cancer cells are employed to specifically target the corresponding cancerous tissue, while white or red blood cells may be used for applications with less specific targets.Most of these cell types were employed to facilitate the precise targeting of nanoparticles to specific tissues.However, some of these cells served a dual purpose by inducing immune stimulation against cancer.
Cervical and ovarian cancer cells were used to favor the cytosolic delivery of cargo inside living cells [21] or for homologous targeting [22].Multiple myeloma cells were chosen to target their equivalent counterparts, ensuring specificity in cargo delivery [25].In the case of melanoma cells, their use was geared towards promoting the delivery and internalization of therapeutic or antigenic materials [12], or for photoimmunotherapy [26].
Leukemia cells were employed to deliver cargo into leukemia cells [34] or were genetically modified to express a protein that can specifically target a tissue [36].Neuroblastoma cells were employed for their capacity to capture neurotoxins effectively [79].Breast cancer cells were used to target homologous cells and deliver cargo [6].Similarly, colon carcinoma [57] head and neck squamous cell carcinoma [58], lung cancer [54], glioma [63,64], glioblastoma [65,66], prostate cancer [67], and liver cancer [68] cells were selected for homologous targeting, ensuring precision in cargo delivery to specific tissues.
In the case of non-cancer cells, leukocytes were harnessed for their capacity to target specific tissues effectively [92].Erythrocytes were used to target cancer tissues, due to their elasticity and capacity to diffuse into the tumor extracellular matrix [81].Dendritic cells were employed to promote tumor immune effects [100].Vaginal endothelial cells were used to protect the cells from a toxin [71].Neural stem cells were used to cross the bloodbrain barrier and specific targeting [72].Neutrophils [101], mesenchymal stem cells [74], fibroblasts [49,69], embryonic kidney cells [70], microglial cells [66], and keratinocytes [73] were also used for specific targeting.
Some investigations opted to combine membranes from different cells so that the coated nanoparticles benefited from the characteristics of both types of source cells.When hybrid membrane-coated nanoparticles were developed by combining two cell types, leukocytes were chosen to mitigate immune recognition [93], platelets were selected for their notable ability to bind to cancer cells [93], and erythrocytes due to their long circulation times [48] and immune-evasion capability [29].Additionally, breast cancer cells [46,48], were incorporated in hybrid membrane coating to ensure precise targeting of homologous cells.

Fragmentation of Cell Membranes
The initial crucial step in the preparation of cell membrane-coated nanoparticles involves the obtention of purified cell membrane fragments.Various techniques are employed to produce these membrane fragments, with hypotonic lysis, homogenization, freeze-thaw, and sonication emerging as the most commonly utilized methods (Figure 2).Often, these methods are used together to enhance results, such as combining hypotonic lysis, homogenization, and freeze-thaw for improved outcomes.In the case of non-cancer cells, leukocytes were harnessed for their capacity to target specific tissues effectively [92].Erythrocytes were used to target cancer tissues, due to their elasticity and capacity to diffuse into the tumor extracellular matrix [81].Dendritic cells were employed to promote tumor immune effects [100].Vaginal endothelial cells were used to protect the cells from a toxin [71].Neural stem cells were used to cross the bloodbrain barrier and specific targeting [72].Neutrophils [101], mesenchymal stem cells [74], fibroblasts [49,69], embryonic kidney cells [70], microglial cells [66], and keratinocytes [73] were also used for specific targeting.
Some investigations opted to combine membranes from different cells so that the coated nanoparticles benefited from the characteristics of both types of source cells.When hybrid membrane-coated nanoparticles were developed by combining two cell types, leukocytes were chosen to mitigate immune recognition [93], platelets were selected for their notable ability to bind to cancer cells [93], and erythrocytes due to their long circulation times [48] and immune-evasion capability [29].Additionally, breast cancer cells [46,48], were incorporated in hybrid membrane coating to ensure precise targeting of homologous cells.

Fragmentation of Cell Membranes
The initial crucial step in the preparation of cell membrane-coated nanoparticles involves the obtention of purified cell membrane fragments.Various techniques are employed to produce these membrane fragments, with hypotonic lysis, homogenization, freeze-thaw, and sonication emerging as the most commonly utilized methods (Figure 2).Often, these methods are used together to enhance results, such as combining hypotonic lysis, homogenization, and freeze-thaw for improved outcomes.

Table 2. Hypotonic lysis buffers used to obtain cell membrane fragments.
Lysis Buffer Used 1

Freeze-Thaw
While not a widely adopted strategy for this purpose, freeze-thaw has been employed in certain experiments [28,37,40,53,59,78,86,93,94,105,106].This technique involves subjecting the cell suspension to multiple cycles of freezing and thawing, with the addition of only a phosphatase inhibitor to the suspension.In some cases, it has been utilized in combination with hypotonic lysis, submitting the lysed cells to several cycles of freezing in liquid nitrogen or at -80 • C and subsequent thawing at 37 • C [21,32,57,63,70].Yao et al. performed a freeze-thaw treatment followed by sonication, without any previous hypotonic lysis [107].

Other Methods
Another method employed for obtaining cell membrane fragments, either used alone or in combination with other techniques, is extrusion.In the studies by Chen et al. and Liu et al. extrusion is applied in conjunction with hypotonic lysis, occurring after the lysis process and before centrifugation to remove other cell components [95,104].

Summary
Upon reviewing all of the compiled articles, hypotonic lysis coupled with homogenization stands out as the overwhelmingly predominant method employed for membrane fragmentation in cells designated for coating.This approach has been consistently applied across a diverse range of cell types, encompassing both cancer and normal cells, and is independent of the specific cell type under investigation.
The hypotonic lysis buffer composed of 20 mM Tris-HCl pH 7.5, 10 mM KCl, 2 mM MgCl 2 , and 1 EDTA-free mini protease inhibitor tablet per 10 mL of solution, emerged as the most prevalent lysis buffer.Remarkably, this buffer was applied across various cell types, including melanoma, myeloma, triple-negative breast cancer, leukemia, and macrophages.Numerous other studies adopted similar lysis buffers based on Tris-HCl, either in combination with other compounds or inhibitors.Nevertheless, buffers incorporating Tris-HCl predominated, demonstrating their widespread usage and satisfactory results.In contrast, homogenization was predominantly carried out using a Dounce homogenizer, underscoring the effectiveness of this device in the membrane fragmentation process.The less commonly employed methods for membrane fragmentation were also ap-plied to various cell types.Freeze-thaw was utilized for the fragmentation of macrophages, melanoma cells, erythrocytes, and platelets, while sonication was applied to cervical cancer, erythrocytes, and macrophages.These findings collectively suggest that there is no singular method universally valid for membrane fragmentation.Instead, there exist several reliable methods for this procedure, irrespective of the cell type chosen for coating.The selection of a specific method appears to be influenced by the availability of required materials in each laboratory.The advantages and disadvantages of each technique are detailed in Table 3.

Membrane Fragments Isolation
After the membranes have been fragmented, the next step involves recovering and isolating these fragments for their subsequent use in coating nanoparticles.Typically, the isolation stage includes 1 to 3 centrifugation steps to separate the remaining membrane materials.This process may be preceded or followed by a gradient separation to move other components away from the membrane fragments.Once the membrane fragments are obtained, they can undergo washing and/or lyophilization, or they may be directly resuspended if the nanoparticle coating process is scheduled immediately after isolation.

Gradient
Certain experiments incorporated a gradient to enhance the performance of membrane fragments between the first and second centrifugations.This gradient took the form of a discontinuous sucrose density gradient, with weight/volume ratios of 55%, 40%, and 30%.The interface between 40% and 30% was then collected [48,92,93].

Other Methods
In two investigations, a lyophilization step was implemented following the centrifugations.Parodi

Summary
In the isolation of membrane fragments, the predominant approach involved subjecting the fragments to one, two, or three centrifugation steps.Some experiments sought to enhance efficiency by incorporating additional steps such as resuspensions in lysis buffer and homogenizations, or by utilizing a sucrose gradient.However, the fundamental procedure typically comprised a combination of one to three centrifugation steps along with the washing of the isolated cell membrane fragments.The optimal number of centrifugations and the inclusion of a gradient appeared to be experiment-specific.While three-step centrifugation with additional lysis and homogenization steps might seem advantageous at first glance, it may not be universally necessary, and in some cases, omitting these extra steps could enhance efficiency.The decision on the specific approach likely depends on the unique requirements and outcomes of each experiment.
The different centrifugation steps and the g forces applied in each are dependent on which cell components are wanted and which ones need to be discarded.Centrifugation around 3000× g served to remove the nuclei and unbroken cells.Centrifugation steps at ca. 10,000× g or 20,000× g are used to remove mitochondria and other organelles.Finally, ultracentrifugation steps are performed to obtain the isolated cell membrane fragments.If the procedure does not require the elimination of organelles, the ultracentrifugation step can be omitted.

Summary
PLGA has emerged as the overwhelmingly preferred material for nanoparticles, primarily due to its notable biocompatibility [22], biodegradability [45] versatile loading capabilities with various cargoes [12].These characteristics make PLGA one of the most suitable materials for nanoparticle coating.The stabilization induced by the coating itself further enhances its utility since both cell membrane fragments and PLGA alone exhibit instability in physiological conditions.However, when united as a coated nanoparticle, their amalgamation remains stable until reaching the target cell.Upon reaching the target cell, the nanoparticle can be released to deliver the cargo effectively [12].
The selection of alternative nanoparticles might hinge on the therapeutic goal.For instance, if phototherapy or radiotherapy is desired, melanin nanoparticles, hollow gold, or copper sulfide nanoparticles may be better suited for the task.The choice of cargo for nanoparticles is entirely dependent on the therapeutic goal.Doxorubicin and dexamethasone stand out as the most frequently employed cargoes, owing to their wellestablished roles in cancer treatments, leveraging their chemotherapeutic [34] and antiinflammatory [35] capabilities, respectively.

Membrane Coating of Nanoparticles
The coating of nanoparticle cores with isolated membrane fragments can be achieved through various methods, with extrusion and sonication being the most common.However, other techniques have also been employed, including a combination of sonication and extrusion.Additionally, in some studies, membrane vesicles were formed before being added to nanoparticles.The ratio of membrane to nanoparticles varies in each experiment, depending on the preceding steps and specific goals of the study.The general protocol for hybrid membrane-coated nanoparticles only derives from the standard on when the coating is applied, as both membrane fragments are mixed (Figure 4).
ever, other techniques have also been employed, including a combination of sonication and extrusion.Additionally, in some studies, membrane vesicles were formed before being added to nanoparticles.The ratio of membrane to nanoparticles varies in each experiment, depending on the preceding steps and specific goals of the study.The general protocol for hybrid membrane-coated nanoparticles only derives from the standard on when the coating is applied, as both membrane fragments are mixed (Figure 4).
Sonication was also employed in cases where membrane vesicles had been previously generated.In these instances, vesicles were sonicated along with nanoparticles in a bath sonicator for 30 s [88], or 2 [82,103], 3 [74], 5 [74], 10 [52] or 40 min [43], at a frequency of 53 kHz and a power of 100 W, or an amplitude of 50%.

Summary
Regarding nanoparticle coating, both sonication and extrusion appear to be valid methods.The frequency with which each method is employed suggests that they yield comparable results.However, a combination of both techniques could potentially enhance efficiency by combining the advantages of each.The advantages and disadvantages of these methods are shown in Table 6.

Discussion
An interesting factor to analyze after reviewing the methods is the membrane isolation efficiency, but almost none of the researchers gave information about it.Zou et al. mentioned how easy the erythrocytes were to isolate [77], while Fang et al. stated that their membrane isolation was successful [12].Only Ferreira et al. gave specific results of the membrane isolation efficiency, reporting that 80% of the membrane was retained after isolation [65].
The coating efficiency is also an important factor to analyze since it shows how successful the coating was.In this regard, most of the researchers report a successful coating, showing the complete coating of the particles with TEM imaging or the analysis of zeta potential comparing the potential of the coated nanoparticles with those of the nude nanoparticle and the isolated membrane.Only 3 of the reviewed articles gave an exact value of coating efficiency.Liu et al. reported a 90.21% efficiency [104], which is in line with the reports of complete or almost complete coating given by all of the investigations that analyzed it with TEM and zeta potential.Conversely, Li et al. report a 21% coating efficiency with a sonication method [50], and Liu et al. measured the coating with a fluorescence quenching essay where they used a quencher that cannot cross membranes and therefore only affects their uncoated parts, state that up to 90% of the nanoparticles are only partially coated and 60% of them are only 20% coated [23].These results open the door for future improvements to the coating techniques.
Most of the coatings caused an increment of around 10 to 30 nm to the diameter of the nanoparticles.But there were many cases where the increase was notably higher, such as .These results can be attributed to an imperfect membrane coating of the nanocarriers, either by having more than one layer of membrane fragments or by the creation of aggregates of those fragments on the surface of the particle.Conversely, Huang et al. report a more exceptional result where they observed a reduction of the size of the nanoparticles, diminishing from 150.1 to 137.3 nm [66].The researches attribute this decrease in size to the pressures to which the particles are subjected to during the extrusion process [66].
The particle-membrane interactions were covered by only a handful of the reviewed articles, since most of them were focused on the effects of the cargo loaded on the nanoparticles on the cells.Despite that, some articles give interesting information about these interactions.Ferreira et al. and Scully et al. explain that the coating is achieved by electrostatic interactions that favor the right-side orientation of the membrane [45,65].Chen et al. and Liu et al. also state that negatively charged nanoparticles give better results than positively charged nanocarriers due to their electrostatic interactions [5,23].Luk et al. stated that the negatively charged cores created a more subtle interaction, allowing the membranes to retain their structure and fluidity, whereas the positively charged cores created strong electrostatic interactions that can cause the collapse of the membrane and thus create aggregates of nanoparticles and membrane fragments [109].Mornet et al. went further and analyzed the effect of differently charged membranes on the coating.They observed that highly negative membranes didn't achieve a successful coating, but moderately negatively charged membranes were able to completely coat the nanoparticles [110].Xia et al. attribute these interactions to the presence of dense negatively charged sialic acid moiety present in the outer membrane side, that allows the right side of the membrane to coat the nanoparticles when a negatively charged core is used but causes the formation of aggregates when positively charged nanoparticles are used due to these negative charges located in the outer side of the membrane [111].Zhao et al. and Zhang et al. state that a higher concentration of H + in the tumoral microenvironment favors the dissociation of the membrane and the nanoparticle, allowing for a faster release of the cargo [24,94].
The biological and micro/nano interactions responsible for tissue-specific therapeutics using these nanoparticles are very diverse.The most common approach was to take profit from the homotypic targeting allowed by the "self-recognition" molecules present on the target tissue [45], especially among those who wanted to target cancers with patient-derived tumor cells, since cancer cells have surface antigens that allow multicellular aggregation through homophilic adhesion domains [100].Some of them rely on the presence of proteins in the membrane coat of the nanocarriers that attach to receptors of the target cells, allowing thus their internalization via endocytosis, such as Tiwari et al. [55], who relied on the presence of heparanase, syndecan-1 and glypican-1, that target HSPG receptors, unchaining the endocytosis.The particles that were designed to avoid immune recognition profited from immune and other blood cells components, especially from macrophages and erythrocytes, respectively, such as macrophages' SIRPα receptor, to which the CD47 proteins of the membranes of the donor cell bind to be recognized by the macrophages and avoid phagocytosis [112].Some opted for the decoration of membranes with targeting molecules, such as aptamers, that target the tumors [78].Another alternative was to genetically modify the donor cells to overexpress a protein that targets a specific protein from the target tissue, such as the rabies viral glycoprotein used by et al. to target acetylcholine receptors on cerebrovascular endothelial cells and nerve cells [72].Another example of this is the use of antibodies linked to the membrane, designed to target the aimed cells [39].
The release kinetics were given by almost all of the reviewed articles, but most of them only studied the difference of released cargo at different pH values.As expected, more cargo was released and also in a faster way when the coated nanocarriers were in more acidic conditions, such as those present at the tumor microenvironments, than in normal physiological conditions (i.e., pH 7.4) [55,78].But among those who actually compared coated and non-coated particles, there were different results.Some researchers such as Qi et al., Zhang et al., Li et al., and Lin et al. report similar release kinetics between both types of carriers, with a minimal difference in speed and total release, as coated nanoparticles were a bit slower and released a bit less cargo than their non-coated counterparts [22,87,91,98].Conversely, Ma et al. observed that coated nanoparticles released less cargo at pH 7.4 but at pH 5.5 were more effective in the release than the non-coated ones [100].Others, such as Li et al. and Chen et al. observed that coated nanoparticles released 10% less of the total cargo than those that were not coated during the first 12-24 h, but in the long term (5-7 days) both end up releasing the same amount of cargo [60,62].Tian et al. observed a great difference in released cargo between coated and non-coated nanoparticles (16.85% against 40.1%), releasing thus less cargo during circulation and improving drug delivery [74].A similar result is reported by Qu et al., who observed a similar difference but both coated and non-coated nanoparticles release higher amounts of cargo (33% versus 50%) [25], and by Scully et al., who reported a 12% release of cargo after 24 h and 16% after 48 h in coated nanocarriers, whereas the non-coated released 30 and 37%, respectively [45].Parodi et al. studied the release kinetics of two different cargos (doxorubicin and BSA) [92].There were very significant differences in the release of both cargos between coated and non-coated carriers, being 20% against 45% release of doxorubicin after 3 h, and 15 versus 25% after 3 h and 80 versus 90% after 48 h, respectively [92].In Liu et al.'s study, non-coated cores were able to deliver the whole cargo after 72 h, but their coated counterparts only released 50% of it in those 72 h, requiring 120 h to release 90% of the cargo [101].Liu stated that the use of PEG and the membrane coating improved the stabilization of the nanoparticles, allowing the reported better retention of the cargo in the nanocarriers [101].Du et al. saw almost no difference in release between coated and non-coated nanocarriers at pH 7.4 (both around 11%) but noticed a significant 16% difference at pH 5.0 [64].Despite not releasing less at physiological conditions and being less efficient at tumor conditions, the low release at pH 7.4 allows for an enhanced cargo accumulation at tumor sites and a reduction of toxicity to other tissues [64].Xie et al. noted that at pH 7.4 coated carriers released much less cargo than the non-coated ones (24.3% against 37.9%), but at pH 5.5, both released more similar amounts (76.1% versus 84.1%) [76].Gong et al. reported a bigger difference at pH 7.4 (40% against 65%), but at pHs 5.5 and 4.7 those differences are reduced significantly, especially at pH 4.7, where the difference is almost negligible [40].These results from Xie et al. and Gong et al. show that the coating protects the nanoparticles and avoids the loss of cargo before arriving at the tumor, improving thus the loading capacity and the drug release behavior [76].
These coating techniques were evaluated through a comparison between cell membranecoated nanoparticles and their non-coated counterparts and/or free cargoes.In all studies conducting cellular uptake analyses, improvements were consistently observed compared to non-coated nanocarriers and free substances.While some studies reported a twofold increase in uptake, others, such as Fang et al., noted a remarkable 40-fold improvement [12].Certain investigations extended their analysis by comparing uptake in the target cell type with other cell types to assess specificity.For instance, Bai et al. observed significantly higher uptake in the target cells compared to other cell types [57].Furthermore, certain studies prioritized investigating immune avoidance, noting a reduction in phagocytosis of coated nanoparticles by macrophages compared to non-coated nanocarriers [38,82,92,101].In summary, cell membrane-coated nanoparticles consistently demonstrated improvements in uptake, specificity, or immune evasion compared to their non-coated counterparts.

Future Directions
While this review primarily concentrates on the cell membrane coating of nanoparticles designed for combating cancers, the application of biomimicry extends beyond oncology.This promising technique has found utility in diverse areas such as gene editing, including the induction of gene expression [31], gene silencing [57], detoxification [79,82], ischemic stroke therapy [101], immune modulation [113], and antibacterial vaccination [36,114,115].The versatility of biomimicry underscores its potential across various fields of research and therapeutic applications.
Further research is needed to enhance the effectiveness of cell membrane-coated nanoparticles, as well as to improve their coating efficiency.While they are already more effective than naked nanoparticles, improvements in targeting ability and residence time are areas of focus.Several novel methods have been explored for this purpose, including modified lipid insertion, membrane hybridization, and genetic modification of source cells [116,117].Modified lipid insertion involves introducing modified lipids into the coated nanoparticles to enhance their fusion and ligand binding properties.For instance, modified lipid insertion has been shown to improve fusion properties [21] and ligand binding properties [84].Membrane hybridization combines characteristics from different source cell membrane fragments [86].For example, creating an erythrocyte-melanoma hybrid coat provides the coated nanoparticles with both the prolonged circulation time of erythrocytes and the homotypic targeting capabilities of melanoma cell membranes [29,46].Genetic modification of source cells involves introducing specific membrane proteins or lipids not present in the original, non-modified cells.This genetic modification provides the coated nanoparticles with the ability to specifically target new ligands, thereby improving their targeting ability [30].

Figure 1 .
Figure 1.Three main steps for obtaining cell membrane-coated nanoparticles: cell lysis and membrane fragmentation, isolation of membrane fragments, and coating selected nanocarriers.The figure has been created with BioRender.com.

Figure 1 .
Figure 1.Three main steps for obtaining cell membrane-coated nanoparticles: cell lysis and membrane fragmentation, isolation of membrane fragments, and coating selected nanocarriers.The figure has been created with BioRender.com.

Figure 2 .
Figure 2. Main strategies used for cell membrane fragmentation: hypotonic lysis; homogenization with probe homogenizer or dounce homogenizer; freeze-thaw and sonication.The figure has been created with BioRender.com.

Figure 2 .
Figure 2. Main strategies used for cell membrane fragmentation: hypotonic lysis; homogenization with probe homogenizer or dounce homogenizer; freeze-thaw and sonication.The figure has been created with BioRender.com.
[87].Jiang et al. and Rao et al. used Hepes B buffer (10 mM Hepes, 5 mM MgCl 2 , 1 mM EDTA, 1 mM DTT, 10 mM KCl, pH 7.6) mixed with protease inhibitor tablets [48,93], and Li et al. used a similar homogenization medium with 20 mM HEPES-NaOH, 1 mM EDTA, and 0.25 M of sucrose with PMSF [22].As an alternative to EDTA-containing buffers, Wang et al. and Park et al., employed EGTA in combination with a phosphatase and protease inhibitor [35,79].Jiang et al. opted for a NaHCO 3 based buffer (1 mM NaHCO 3 , 0.2 mM EDTA•2Na, 1 mM PMSF and 1 × PIC in H 2 O) [39], while Du et al. used a similar buffer [64].Li et al. used double distilled water [103].Some articles did not specify the exact buffer composition but indicated the use of a low-osmotic lysis buffer containing membrane protein extraction reagents and PMSF [26].Wu et al. subjected the cell mix to only a membrane protein extraction buffer et al. lyophilized the isolated membranes before rehydrating them and storing them at 4 • C [92].On the other hand, Bai et al. and Nie et al. directly lyophilized the membranes and stored them at −80 • C for future use [6,40,57,99].

Figure 4 .
Figure 4. Scheme of the preparation of hybrid membrane-coated immunomagnetic beads (HM-IMBs) for high-performance isolation of circulating tumor cells (CTCs).(a) Platelet (PLT) and leukocyte (WBC) membranes, along with their associated proteins, were independently separated from blood samples, fused, and then coated onto MBs.Then, the resulting PLT-WBC HM-coated MBs were surface-modified with antibodies to form HM-coated immuno-MBs.(b) HM-IMBs inherited

Figure 4 .
Figure 4. Scheme of the preparation of hybrid membrane-coated immunomagnetic beads (HM-IMBs) for high-performance isolation of circulating tumor cells (CTCs).(a) Platelet (PLT) and leukocyte (WBC) membranes, along with their associated proteins, were independently separated from blood samples, fused, and then coated onto MBs.Then, the resulting PLT-WBC HM-coated MBs were surface-modified with antibodies to form HM-coated immuno-MBs.(b) HM-IMBs inherited enhanced CTCs binding from PLTs and the property of reduced interaction with homologous WBCs from WBCs, was used for high-efficiency and high-purity isolation of CTCs.Copy from Ref. [93], Wiley © 2018.

Table 1 .
Donor cell types for nanoparticle coating applications.

Table 3 .
Advantages and disadvantages of the membrane fragmentation techniques.

Table 4 .
Nanoparticles used for membrane coating.

Table 5 .
Cargoes loaded in the nanoparticles.

Table 6 .
Advantages and disadvantages of membrane coating techniques.