Freezing and sublimation effects on cryo-SEM imaging and microanalysis

Liang, Jenn‐Tai; Xiao, Xiong; Chou, Tsengming; Libera, Matthew

doi:10.1017/s1431927619006275

Cited by 8 publications

(10 citation statements)

References 5 publications

(5 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In this investigation, the cryo-SEM technique was also performed in addition to conventional SEM, to visualize M. chimaera biofilm in a form closer to its native state with the ultra-structures and cell components preserved within the biofilm. It is widely reported in the literature that preparation steps for conventional FE-SEM can compromise the structural integrity of the native state of biofilm ( Wu et al, 2014 ; Liang et al, 2019 ). For example, the dehydration step during sample preparation can lead to distortion of the bacterial extracellular matrix and can introduce dramatic artifacts ( Azeredo et al, 2017 ; Liang et al, 2019 ).…”

Section: Discussionmentioning

confidence: 99%

“…It is widely reported in the literature that preparation steps for conventional FE-SEM can compromise the structural integrity of the native state of biofilm ( Wu et al, 2014 ; Liang et al, 2019 ). For example, the dehydration step during sample preparation can lead to distortion of the bacterial extracellular matrix and can introduce dramatic artifacts ( Azeredo et al, 2017 ; Liang et al, 2019 ). The cryo-SEM images revealed surface ultrastructure and fiber-like projections extending between cells in early M. chimaera biofilms of 24 h and throughout the biofilm by week 2 ( Figures 4 , 5 ).…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Characterization of Biofilm Formation by Mycobacterium chimaera on Medical Device Materials

et al. 2021

View full text Add to dashboard Cite

Non-tuberculous mycobacteria (NTM) are widespread in the environment and are a public health concern due to their resistance to antimicrobial agents. The colonization of surgical heater-cooler devices (HCDs) by the slow-growing NTM species Mycobacterium chimaera has recently been linked to multiple invasive infections in patients worldwide. The resistance of M. chimaera to antimicrobials may be aided by a protective biofilm matrix of extracellular polymeric substances (EPS). This study explored the hypothesis that M. chimaera can form biofilms on medically relevant materials. Several M. chimaera strains, including two HCD isolates, were used to inoculate a panel of medical device materials. M. chimaera colonization of the surfaces was monitored for 6 weeks. M. chimaera formed a robust biofilm at the air-liquid interface of borosilicate glass tubes, which increased in mass over time. M. chimaera was observed by 3D Laser Scanning Microscopy to have motility during colonization, and form biofilms on stainless steel, titanium, silicone and polystyrene surfaces during the first week of inoculation. Scanning electron microscopy (SEM) of M. chimaera biofilms after 4 weeks of inoculation showed that M. chimaera cells were enclosed entirely in extracellular material, while cryo-preserved SEM samples further revealed that an ultrastructural component of the EPS matrix was a tangled mesh of 3D fiber-like projections connecting cells. Considering that slow-growing M. chimaera typically has culture times on the order of weeks, the microscopically observed ability to rapidly colonize stainless steel and titanium surfaces in as little as 24 h after inoculation is uncharacteristic. The insights that this study provides into M. chimaera colonization and biofilm formation of medical device materials are a significant advance in our fundamental understanding of M. chimaera surface interactions and have important implications for research into novel antimicrobial materials, designs and other approaches to help reduce the risk of infection.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Characterization of Biofilm Formation by Mycobacterium chimaera on Medical Device Materials

et al. 2021

View full text Add to dashboard Cite

show abstract

“…The samples were then sputtered with 5 nm gold before imaging by SEM (TESCAN Mira3) at room temperature. The microspheres were also suspended in phosphate buffered saline (PBS) buffer and then were cryo-fixed with a high-pressure freezer (Leica ICE) to preserve their native structures [3,4]. Cross-sectional specimens were prepared by freeze-fracture (Leica ACE 600).…”

Section: Methodsmentioning

confidence: 99%

Micro-imaging and Microanalysis of PLGA Complex Drug Products by Scanning Electron Microscopy (SEM)

Liang¹,

Wang²,

Qin³

et al. 2020

Microsc Microanal

Self Cite

View full text Add to dashboard Cite

“…Therefore, there is no need to freeze-etch samples like BPV-MVLs, which have enough imaging contrast without any sublimation. For other samples with high water content and no sharp interface (e.g., hydrogels, biological samples), sublimation could be crucial for satisfactory imaging (Liang et al, 2019b).…”

Section: Of 17mentioning

confidence: 99%

Characterization of Complex Drug Formulations Using Cryogenic Scanning Electron Microscopy (Cryo‐SEM)

Liang

Koo

et al. 2022

Current Protocols

Self Cite

View full text Add to dashboard Cite

The physicochemical properties of complex drug formulations, including liposomes, suspensions, and emulsions, are important for understanding drug release mechanisms, quality control, and regulatory assessment. It is ideal to characterize these complex drug formulations in their native hydrated state. This article describes the characterization of complex drug formulations in a frozen‐hydrated state using cryogenic scanning electron microscopy (cryo‐SEM). In comparison to other techniques, such as optical microscopy or room‐temperature scanning electron microscopy, cryo‐SEM combines the advantage of studying hydrated samples with high‐resolution imaging capability. Detailed information regarding cryo‐fixation, cryo‐fracture, freeze‐etching, sputter‐coating, and cryo‐SEM imaging is included in this article. A multivesicular liposomal complex drug formulation is used to illustrate the impact of different cryogenic sample preparation conditions. In addition to drug formulations, this approach can also be applied to biological samples (e.g., cells, bacteria) and soft‐matter samples (e.g., hydrogels). © Published 2022. This article is a U.S. Government work and is in the public domain in the USA. Basic Protocol 1: Cryo‐fixation to preserve the native structure of samples using planchettes Alternate Protocol: Cryo‐fixation to preserve the native structure of biological samples on sapphire disks Basic Protocol 2: Sample preparation for cross‐sectional cryo‐SEM imaging Basic Protocol 3: Cryo‐SEM imaging and microanalysis

show abstract

Freezing and sublimation effects on cryo-SEM imaging and microanalysis

Cited by 8 publications

References 5 publications

Characterization of Biofilm Formation by Mycobacterium chimaera on Medical Device Materials

Characterization of Biofilm Formation by Mycobacterium chimaera on Medical Device Materials

Micro-imaging and Microanalysis of PLGA Complex Drug Products by Scanning Electron Microscopy (SEM)

Characterization of Complex Drug Formulations Using Cryogenic Scanning Electron Microscopy (Cryo‐SEM)

Contact Info

Product

Resources

About