A simple method for exposing and examining the interior of fragile biological materials

1990 ◽  
Vol 48 (3) ◽  
pp. 858-859
Author(s):  
Arthur J. Wasserman ◽  
Y. Pedro Kato ◽  
Frederick H. Silver
Keyword(s):  
Liquid Nitrogen ◽  
Mechanical Damage ◽  
Aluminum Foil ◽  
Collagen Fibers ◽  
Fiber Formation ◽  
Bottom Surface ◽  
Electron Microscopic ◽  
Simple Method ◽  
Bovine Hide ◽  
Polyethylene Tubing

Examining the inside of delicate and fragile dry biomaterials is difficult because they are vulnerable to mechanical damage. Compression and sheering of a sample during exposure of the interior can produce artifacts making interpretation of the ultrastructure difficult. In this report a simple method for exposing and retaining the interior substructure of delicate specimens and mounting them for scanning electron microscopic (SEM) observation is described.Collagen fibers were prepared as described previously. In brief, 1% collagen dispersion, prepared from bovine hide, was extruded through polyethylene tubing with an inner diameter of 0.28 mm into a 37°C, pH 7.5, fiber formation buffer. After 45 mins in the buffer, the fibers were rinsed in isopropyl alcohol for 4 hrs followed by distilled water for 20 mins. The fibers were then crosslinked.The interior of collagen fibers were exposed by deep freezing 1 cm segments of fiber in liquid nitrogen. Using iris scissors the first and last piece of each segment (which contained ends previously exposed to the atmosphere) were snapped away and discarded. Each frozen segment was then snapped in half. By this method each half of the original 1 cm segment had a freshly cleaved top and bottom surface. The segments were transferred to an aluminum foil pouch (in liquid nitrogen).

Methionine-Specific Staining of Collagen

1982 ◽  
Vol 40 ◽  
pp. 294-295
Author(s):  
E.M. Kuhn ◽  
K.D. Marenus ◽  
M. Beer
Keyword(s):  
Amino Acids ◽  
Collagen Fibers ◽  
Type Iii Collagen ◽  
Type I ◽  
Electron Microscopic ◽  
Good Correspondence ◽  
Specific Staining ◽  
Type Iii ◽  
Different Types ◽  
Sulfur Containing

Fibers composed of different types of collagen cannot be differentiated by conventional electron microscopic stains. We are developing staining procedures aimed at identifying collagen fibers of different types.Pt(Gly-L-Met)Cl binds specifically to sulfur-containing amino acids. Different collagens have methionine (met) residues at somewhat different positions. A good correspondence has been reported between known met positions and Pt(GLM) bands in rat Type I SLS (collagen aggregates in which molecules lie adjacent to each other in exact register). We have confirmed this relationship in Type III collagen SLS (Fig. 1).

scholarly journals In vitro Conservation and Cryopreservation of Nandina domestica, an Outdoor Ornamental Shrub

2013 ◽  
Vol 41 (2) ◽  
pp. 638 ◽  
Author(s):  
Aylin OZUDOGRU ◽  
Diogo Pedrosa Corrêa Da SILVA ◽  
Ergun KAYA ◽  
Giuliano DRADI ◽  
Renato PAIVA ◽  
...  
Keyword(s):  
Liquid Nitrogen ◽  
Storage Temperature ◽  
Sucrose Concentration ◽  
Aluminum Foil ◽  
Shoot Tips ◽  
In Vitro Conservation ◽  
Shoot Cultures ◽  
Storage Medium ◽  
Droplet Vitrification

The study focused on an economically-important ornamental outdoor shrub, Nandina domestica, with the aims to (i) optimize an effective in vitro conservation method, and (ii) develop a cryopreservation protocol for shoot tips by the PVS2 vitrification and droplet-vitrification techniques. For in vitro conservation of shoot cultures, the tested parameters were sucrose content in the storage medium (30, 45, 60 g/L) and storage temperature (4 °C or 8 °C). Cryopreservation was performed by applying the PVS2 vitrification solution, in 2-ml cryovials or in drops over aluminum foil strips, for 15, 30, 60 or 90 min at 0 °C, followed by the direct immersion in liquid nitrogen of shoot tips. Results show that N. domestica shoots can be conserved successfully for 6 months at both the temperatures tested, especially when 60 g/L sucrose is used in the storage medium. However, conservation at 4 °C showed to be more appropriate, as hyperhydricity was observed in post-conservation of shoots coming from storage at 8 °C. As for cryopreservation, a daily gradual increase of sucrose concentration (from 0.25 to 1.0 M) produced better protection to the samples that were stored in liquid nitrogen. Indeed, with this sucrose treatment method, a 30-min PVS2 incubation time was enough to produce, 60 days after thawing, the best recovery (47% and 50%) of shoot tips, cryopreserved with PVS2 vitrification and droplet-vitrification, respectively.

A Very Low-Cost, Labor-Efficient, and Simple Method to Block Scattered Ultraviolet Light in PDMS Microfluidic Devices by Inserting Aluminum Foil Strips

2018 ◽  
Vol 11 (1) ◽  
Author(s):  
Shuo Wang ◽  
Peter Shankles ◽  
Scott Retterer ◽  
Yong Tae Kang ◽  
Chang Kyoung Choi
Keyword(s):  
Soft Lithography ◽  
Uv Light ◽  
Low Cost ◽  
Microfluidic Devices ◽  
Aluminum Foil ◽  
Material Synthesis ◽  
Simple Method ◽  
Micro Particles ◽  
Complex Shapes ◽  
The Cost

Abstract Opto-microfluidic methods have advantages for manufacturing complex shapes or structures of micro particles/hydrogels. Most of these microfluidic devices are made of polydimethylsiloxane (PDMS) by soft lithography because of its flexibility of designing and manufacturing. However, PDMS scatters ultraviolet (UV) light, which polymerizes the photocrosslinkable materials at undesirable locations and clogs the microfluidic devices. A fluorescent dye has previously been employed to absorb the scattered UV light and shift its wavelength to effectively solve this issue. However, this method is limited due to the cost of the materials (tens of dollars per microchip), the time consumed on synthesizing the fluorescent material and verifying its quality (two to three days). More importantly, significant expertise on material synthesis and characterization is required for users of the opto-microfluidic technique. The cost of preliminary testing on multiple iterations of different microfluidic chip designs would also be excessive. Alternatively, with a delicate microchannel design, we simply inserted aluminum foil strips (AFS) inside the PDMS device to block the scattered UV light. By using this method, the UV light was limited to the exposure region so that the opto-microfluidic device could consistently generate microgels longer than 6 h. This is a nearly cost- and labor-free method to solve this issue.

scholarly journals Cryo-shocked cancer cells for targeted drug delivery and vaccination

2020 ◽  
Vol 6 (50) ◽  
pp. eabc3013
Author(s):  
Tianyuan Ci ◽  
Hongjun Li ◽  
Guojun Chen ◽  
Zejun Wang ◽  
Jinqiang Wang ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Cancer Cells ◽  
Liquid Nitrogen ◽  
Targeted Drug Delivery ◽  
Targeted Delivery ◽  
Myeloid Leukemia ◽  
Live Cells ◽  
Simple Method ◽  
Drug Delivery Vehicles ◽  
Delivery Vehicles

Live cells have been vastly engineered into drug delivery vehicles to leverage their targeting capability and cargo release behavior. Here, we describe a simple method to obtain therapeutics-containing “dead cells” by shocking live cancer cells in liquid nitrogen to eliminate pathogenicity while preserving their major structure and chemotaxis toward the lesion site. In an acute myeloid leukemia (AML) mouse model, we demonstrated that the liquid nitrogen–treated AML cells (LNT cells) can augment targeted delivery of doxorubicin (DOX) toward the bone marrow. Moreover, LNT cells serve as a cancer vaccine and promote antitumor immune responses that prolong the survival of tumor-bearing mice. Preimmunization with LNT cells along with an adjuvant also protected healthy mice from AML cell challenge.

A simple method of rapid freezing adequately preserves brain tissue for immunocytochemistry, light and electron microscopic examination

1993 ◽  
Vol 86 (6) ◽  
pp. 645-650 ◽  
Author(s):  
David Nochlin ◽  
Alan P. Mackenzie ◽  
Eileen M. Bryant ◽  
Thomas H. Norwood ◽  
S. Mark Sumi
Keyword(s):  
Brain Tissue ◽  
Microscopic Examination ◽  
Electron Microscopic Examination ◽  
Rapid Freezing ◽  
Electron Microscopic ◽  
Simple Method

209 VIABILITY OF PORCINE EMBRYOS VITRIFIED BY THE METAL MESH VITRIFICATION METHOD AFTER SURGICAL OR NONSURGICAL TRANSFER

2007 ◽  
Vol 19 (1) ◽  
pp. 221 ◽  
Author(s):  
Y. Fujino ◽  
Y. Nakamura ◽  
H. Kobayashi ◽  
S. Nakano ◽  
C. Suzuki ◽  
...  
Keyword(s):  
Liquid Nitrogen ◽  
Uterine Horn ◽  
Outer Diameter ◽  
Metal Mesh ◽  
Angle Bend ◽  
Fetal Bovine ◽  
Inner Diameter ◽  
Polyethylene Tubing ◽  
Practical Measure ◽  
Uterine Body

The aim of the present study was to evaluate viability of porcine embryos vitrified by the metal mesh vitrification (MMV) method after surgical or nonsurgical transfer. Prepubertal gilts were treated with eCG and hCG (= Day 0), and then inseminated artificially. Expanding blastocysts that were about 200 �m in diameter were collected on Day 7. The embryos were equilibrated in 7.5% ethylene glycol (EG) + 7.5% DMSO + 20% fetal bovine serum (FBS) in PBS at 37�C for 5 min, and then transferred into 15% EG + 15% DMSO + 0.6 M trehalose + 20% FBS in PBS for 1 min. Embryos in groups of 5 were transferred in a minimum volume of the vitrification solution (less than 1 �L) onto stainless steel mesh (75 �m screen size, 1.5 mm in width by 10 mm in length, with a 3-mm right-angle bend), and then plunged into liquid nitrogen. The mesh was stored in a 1.8-mL cryotube submerged in liquid nitrogen. Warming and dilution were performed by moving the mesh from liquid nitrogen into 0.5 M trehalose + 20% FBS in PBS at 37�C for 5 min. Embryos were rinsed twice in NCSU37 + 10% FBS (mNCSU37) for 5 min. After being vitrified, embryos in groups of 20 per recipient were suspended in modified NCSU37 medium and then transferred into gilts either by surgical transfer (5 gilts) or by nonsurgical transfer (6 sows). For surgical transfer, embryos suspended in 0.1 mL of medium were transferred into the uterine horn at 15 cm above the uterine body, which was about 35 cm from the external uterine orifice. For nonsurgical transfer, an intrauterine catheter made from polyethylene tubing (1.2 m long, 3.0 mm outer diameter, 0.5 mm inner diameter) was used. A spiral guide inserted through the vagina into the cervix was used to guide the catheter into one uterine horn. The catheter was moved through the cervix and along the uterine horn. Then, embryos suspended in 1 mL of medium were transferred. Pregnancy was assessed by ultrasonography at 30 days post-estrus. With surgical transfer, 4 of 5 recipients became pregnant, and 3 gilts farrowed a total of 21 (10, 8, 3) live piglets; the fourth gilt aborted one fetus on Day 34. With nonsurgical transfer, 3 of 6 sows became pregnant. The present study demonstrates that vitrified porcine embryos can develop after both surgical and nonsurgical transfer to recipients. As a practical measure, nonsurgical transfer of vitrified porcine blastocysts may be used instead of surgical transfer.

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