이 연구는 농업과 농식품 캐나다 -베이스 프로젝트 &quot;마커를 이용한 선택의 변수 품질 특성에 대한 노믹 격차를 줄이기 위해 고기 샘플에 대한 높은 처리량 기술 개발&quot;의 일환이었다. 쇠고기 장치와 라콤 연구 센터에서 육류 가공 직원의 숙련 된 지원을 진심으로 감사합니다. 저자는 또한 크리스틴 Burbidge - 보이드, 프랜 코스텔로, 글리 니스 Croken 및 Rhona 대커의 전용 기술 지원을 감사드립니다.

1. 스테이크 컬렉션 <ol><li> 24 시간에서 다음 등급 - 48 시간 후 도축, longissimus 근육 (중 흉부 및 / 또는 lumborum 끝) 쇠고기 시체에서 제거 및 품질 분석을 위해 수집됩니다. </li><li> 근육 피하 지방과 정사각형의 정돈되어 있습니다. 근육 다음 안전 및 식품 처리 프로토콜에 따라 2.5 cm (1) 스테이크로 잘라된다. 개별 스테이크의 절단에 따라 적절한 라벨 (동물 번호, 사이드, 근육의 위치, 또는 시간) 각각의 스테이크에 배치됩니다. </li><li> 스테이크가 다음 저장을 위해 준비가되어 있습니다. 그들은 신선한 유지하는 경우, 그들은 트레이에 배치하고 그릴로 이동합니다. 신선한 스테이크 바로 조리 할 수​​없는 경우, 그들은 요리 탈수를 방지하기 시작할 수있을 때까지 플라스틱으로 덮여해야합니다. 스테이크 쿨러 세, 또는 냉동 할 경우, 무게는 산소 투과성 진공 포장 BA에 배치되기 전에 두 개의 소수 자릿수를 기록G와 밀봉은 진공 포장 기계를 사용하여. 스테이크 그런 다음 각각의 장기 저장 위치에 배치됩니다. </li></ol> 2. 조리 과정 <ol><li> 시료가 동결 된 경우, 하룻밤 해동하는 4 ° C 쿨러의 냉장고와 장소에서 제거합니다. </li><li> 요리의 날에, 210 예열 오픈 난로 그릴 ° C 샘플 조리 및 스캔 장치의 출구에 열전대를 삽입하여 온도 로거 시스템과 컴퓨터를 준비합니다. 살짝 기름 요리가 시작되기 전에 야채 약 10 분 단축에 그릴. </li><li> 요리를 시작하기 전에, 쿨러에서 스테이크를 제거합니다. 패키지에서 샘플을 제거하고 종이 타월로 스테이크에서 초과 수분을 얼룩. 소수점 이하 두 자리에 스테이크의 원시 무게를 기록합니다. </li><li> 의 길이 방향 축을 따라 스테이크의 중간에 열전대를 삽입합니다. 조리 데이터를 기록 열전대 스캔을 시작합니다. </li><li> 배치그릴에 스테이크는 스테이크와 그릴의 평평한 표면을 요리에 열전대 평행을 유지. 프로브가 평행 유지되지 않는 경우, 부적절한 온도 측정 될 수 있습니다. </li><li> 35.5의 내부 온도에 스테이크를 요리 ° C 다음, 샘플을 뒤집어 71의 최종 내부 온도에 요리를 계속 ° C. 내부 온도가 71 ° C가되면, 그릴에서 샘플을 제거하고 스테이크의 열전대를 잡아 당깁니다. </li><li> 요리 중량이 요구되는 경우, 약간 냉각 스테이크 (1-2 분)은, 다음 과잉 수분을 제거하고 소수 두 자리의 샘플의 무게를 위해 종이 타월로 샘플을 가릴 수 있습니다. </li><li> 조리 데이터가 필요한 경우, 열전대 스캔 데이터를 저장합니다. </li></ol> 3. 각도 조정 및 샘플 준비 <ol><li> 정사각형 끝 부분을 만들 수있는 스테이크의 측면 끝에서 2cm - 샘플은 여전히​​ 그릴에서 따뜻한이지만, 컷 1 - 1. </li><li> WID에 걸쳐 5cm 섹션스테이크 번째 병렬 원래 컷에, 5cm에서 칼 노치 &#39;샘플 크기 조정 상자&#39;를 사용하여 준비가되어 있습니다. </li><li> 근육 섬유 각도의 방향은 스테이크 나이프 컷 표면에 각도기를 배치하여 측정 할 수 있습니다. </li><li> 가변 각도 절단 상자는 다음 두 블레이드 칼을 안내하는 바닥 구멍을 찾을 하부 금속판을 슬라이딩하여 샘플의 해당 각도로 설정되어 있습니다. 해당 각도 낮은 금속판을 고정 노브를 조입니다. </li><li> 근육 섬유 방향이 더블 블레이드 칼의 각도와 평행이되도​​록 가변 각도 절단 상자의 절단 영역에 스테이크 5cm 섹션을 배치합니다. </li><li> 더블 블레이드 칼 칼 해당 각도에서 전체 샘플을 통과 할 수 있도록, 그래서 이전에 잘라 5cm 조각에 걸쳐 1cm 슬라이스를 잘라하는 데 사용됩니다. 스테이크를 배치 할 때, 그것은 가장 가까운 절삭 영역의 가장자리를 만지고 있는지 확인사용자는 절단 모션으로 사용자를 향해 스테이크를 끌어 것입니다. </li><li> 절단, 절단 상자에 이중 블레이드 칼을 넣고 1cm 지구로 고기를 잘라 칼을 잡아 당깁니다. 사용자를 향해 칼을 당기면서 고기를 찢어 방지하기 위해 약간의 톱질 동작을 사용합니다. 가이드 판의 한쪽 가장자리에 칼날이 동일 절단 비율을 유지하기 위해 계속. </li><li> 결과는 섬유가 지속적으로 슬라이스 평행으로, 5cm 길이의 따뜻한 1cm 두께의 슬라이스 여야합니다. 이 조각은 그 고기 질감의 조각 전단력 분석을 위해 사용됩니다. </li></ol> 4. 전단력 분석 <ol><li> 전단력 분석을 확인하려면, 슬라이스 전단력 프로토콜 (SSF)는 가변 각도 절단 상자를 얻을 슬라이스 5cm가 1cm를 사용합니다. 평면 전단 슬라이스 블레이드 텍스처 분석기 기계를 사용하여 분석 따뜻한 슬라이스에서 수행됩니다. </li><li> 50kg로드 셀 세트 텍스처 분석기를 준비500mm / min의 크로스 헤드 속도를 가진 48mm에 전단 거리입니다. </li><li> 전단은 전단 슬라이스 섬유에 필요한 최대 힘의 정확한 분석을 제공하기 위해 근육 섬유에 수직 한 번 수행됩니다. </li><li> 슬라이스 센터를 통해 완전히 전단되면 저장하고 데이터를 기록합니다. </li></ol>

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우리는 공개 아무것도 없어.

각도 조절 상자 SSF 블레이드는 항상 오히려 고정 45 ° 각도로 잘라 8에서 일어날 수있는 약 직각보다 진정한 수직 각도로 근육 섬유를 가위 보장합니다. 가변 각도 절단 상자의 응용 프로그램은 샘플의 최대 전단력보다 정확한 묘사를 허용, 그것은 가변 각도 절단 상자가 개발 한 객관적인 품질 분석 기법의 개선이 기초에 있습니다. 또한, 각도 조절 박스 ​​(30 ° ~ 60 °)에 의해 제시 각도?...

부드러움 고기 1에서 가장 중요한 품질 속성 중 하나입니다. 쇠고기 부드러움의 불일치 쇠고기 산업이 직면하고있는 주요 문제 중 하나로 확인되었습니다. 그것은 틀림없이이 최고를 보여 주었다 수단 방법으로 정확하게 기계로 가공 전단 플레이트 삼각형 구멍이 특징 워너 Bratzler 전단력 (WBSF) 시험, 고기 감각 부드러움 3,4를 나타내는 데 사용되는 가장 널리 방법 고기 인성 5 감각 패널 점수 상관 관계. 그러나, 슬라이스 전단력 프로토콜 (SSF)는 표준 WBSF 프로토콜 7에 대한 대안으로, 근육의 질감과 부드러움 6 분석을위한 중요한 기술이되고있다. 그것은 신속한 분석 또는 샘플의 높은 숫자를 처리해야하는 경우에서 유용합니다. 여전히 따뜻하면 SSF를 들어, 하나의 코어는 스테이크에서 가져온 것입니다, 대 멀티 코어 (3-6) 일반적으로 선미 스테이크에서 촬영WBSF 6 냉장 어 24 시간. 그것은 부드러움의 평균 WBSF에 가장 적합한 표현을하지 스테이크의 중심과 중앙에, 스테이크 9의 외측 내측면 사이에 차이가 발견 되었기 때문에이 한 조각에서, SSF는 전체 스테이크 (8)의 평균 텍스처를 분석 . SSF의 몰락 스테이크 내 옆에서 내측 측면에, 전단력 값 9의 변화가 그이지만, 지속적으로 5cm 섹션을 만들기 위해 크기 조정 상자를 사용하여, 그것은 올 수 변동을 줄일 수 있습니다 스테이크의 다른 섹션을 사용. 어떤 스테이크가 서로 다른 크기 때문에 그러나 5 센티미터 슬라이스는 전단력 값이 10에 영향을 미칠 수있는 스테이크의 다른 위치에서 발생할 수 있습니다. 반면에, SSF 프로토콜을 분석 할 수있는 부분을 얻을 수 있도록 설계 원래 절단 상자는 2 고정 각도 45 °와 90 °의, 그러나, 근육스테이크 내에서, 그리고 근육의 11에서 섬유 방향 변경. SHACKELFORD 및 휠러 (12)는 평균 longissimus 각도는 43.8 가까운 45 ℃에서, ° 및 샘플이 항상 같은 위치에서 수집되었을 때 따라서 적절하다고 인정하는 것을 밝혔다. 그러나 Derington 등은. 11 33.1 °, 53.9 ° 사이의 longissimus 근육을 따라 섬유의 각도 범위를보고했다. 따라서, 때, 연구 목적, 같은 동물에서 여러 스테이크 45 ° 또는 90 ° 잠재적 정확성을 감소 하나에 모든 섬유 각도를 지정, 분석 할 필요가있다. 절단 각도를 측정하고 조정하는 능력을 갖는 것은 슬라이스 지속적으로 근육 섬유와 병렬로 실행하도록 허용하는 가변 각도 절단 상자의 사용, 최대 전단력보다 정확한 묘사를 제공 할 수 있습니다.

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	<tbody>
		<tr>
			<td>Name of equipment</td>
			<td>Company</td>
			<td>Catalogue number</td>
		</tr>
		<tr>
			<td>Vacuum Packager</td>
			<td>Koch Equipment, Kansas City, MO, USA</td>
			<td>Model UV2100</td>
		</tr>
		<tr>
			<td>Vacuum bags</td>
			<td>Winpak Ltd., Winnipeg, MB, Canada</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Thermocouples and Scanning Device</td>
			<td>Agilent / Hewlett Packard, Santa Clara, CA, USA</td>
			<td>34970A Data Acquisition Switch Unit</td>
		</tr>
		<tr>
			<td>Grill</td>
			<td>Garland Commercial Ranges Ltd., Mississauga, ON, Canada</td>
			<td>Model ED-30-B</td>
		</tr>
		<tr>
			<td>Crisco Vegetable Shortening</td>
			<td>The J.M. Smucker Company, Orrville, OH, USA</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Sample sizing box</td>
			<td>G-R Manufacturing Co., Manhattan, KS, USA</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Angle adjustable box</td>
			<td>Innovation Centre, Red Deer College, Red Deer, AB, Canada</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Texture Analyzer Machine</td>
			<td>Texture Technologies, Hamilton, MA, USA</td>
			<td>Model TA-XT Plus</td>
		</tr>
		<tr>
			<td>Load Cell, 50 kg</td>
			<td>Texture Technologies, Hamilton, MA, USA</td>
			<td>TA-XT Plus</td>
		</tr>
		<tr>
			<td>USDA Warner-Bratzler knife w/guillotine block</td>
			<td>Texture Technologies, Hamilton, MA, USA</td>
			<td>TA-7</td>
		</tr>
		<tr>
			<td>Flat rectangle Blade for Slice Shear Force</td>
			<td>Texture Technologies, Hamilton, MA, USA</td>
			<td>TA-7C</td>
		</tr>
	</tbody>
</table>

introducing an angle adjustable cutting box for analyzing slice shear force in meat

연구 longissimus 근육의 섬유 각도 스테이크 내부와 근육을 통해 위치에 따라 달라질 수 있음을 나타냅니다. 대신 원래 고정 45 ° 또는 90 ° 전단력 시험을위한 절단 각도를 사용하는 칼의 각도는 각 샘플의 섬유 각도에 해당하므로, 가변 각도 절단 상자를 조정할 수 있습니다. 2 분 안에 71의 내부 온도로 조리 후 ° 210 세트 오픈 난로 그릴에 C ° C 5 센티미터 코어에서 1cm가 2 나이프 블레이드를 사용하여, 근육 섬유 방향에 평행하게, 스테이크에서 절단 세트 1 떨어져 CM. 이 따뜻한 코어는 다음 고기 질감을 평가하는 슬라이스 전단력 프로토콜 (SSF)를 받게됩니다. 가변 각도 절단 상자와 SSF 프로토콜의 사용은 조각과 근육 섬유가 지속적으로 평행으로, 최대 전단력의 정확한 표현을 제공합니다. 따라서, 가변 각도 절단 상자, SSF 프로토콜과 함께, 높은 번째로 사용할 수 있습니다roughput 기술은 정확하게 다른 근육에 잠재적 longissimus 근육의 다른 위치에 고기 부드러움을 평가하고 있습니다.

31 개의 완성 된 상업 쇠고기 소들이 도살되었고, 그 시체는 (5백28~6백1kg) 분리 하였다. 왼쪽 측면은 2 ℃에서 후 3 시간에 대해 10 ° C에서 개최하는 동안 부드러움의 변화를 만들기 위해, 오른쪽은 즉시, 2 ° C에서 보관 하였다 오른쪽과 왼쪽 측면에서 longissimus 근육은 도축 후 24 시간을 제거 하였다. 왼쪽에서 샘플 ( &quot;TENDER&quot;처리) 분석되기 전에 6 일 동안 세 동안 오른쪽 ( &quot;TOUGH&quot;처...

Watch this Scientific Journal Video about Introducing an Angle Adjustable Cutting Box for Analyzing Slice Shear Force in Meat at JoVE.com

Introducing an Angle Adjustable Cutting Box for Analyzing Slice Shear Force in Meat

슬라이스 전단력 쇠고기 텍스처 분석을위한 참조 방법입니다. 각도 조절 절단 상자를 사용하여 연구 목적에 대한 정확도를 높일 수 있습니다. 내의 다른 위치에서의 결과<em&gt; longissimus</em&gt; 근육 워너 Bratzler 전단력 방법론과 다른 근육 가능성이 높은 적응력과 높은 상관 관계를 보여줍니다.

고기를 슬라이스 전단력 분석을위한 각도 조절 절단 상자 소개

Research indicates the fibre angle of the longissimus muscle can vary, depending upon location within a steak and throughout the muscle. Instead of using ...

introducing-an-angle-adjustable-cutting-box-for-analyzing-slice-shear

Research

JoVE Journal

Biology

12.8K 조회수.  Agriculture and Agri-Food Canada. 슬라이스 전단력 쇠고기 텍스처 분석을위한 참조 방법입니다. 각도 조절 절단 상자를 사용하여 연구 목적에 대한 정확도를 높일 수 있습니다. 내의 다른 위치에서의 결과 longissimus 근육 워너 Bratzler 전단력 방법론과 다른 근육 가능성이 높은 적응력과 높은 상관 관계를 보여줍니다.

비디오: Introducing an Angle Adjustable Cutting Box for Analyzing Slice Shear Force in Meat

Organotypic Slice Culture of E18 Rat Brains

Organotypic slice cultures from embryonic rodent brains are widely used to study brain development. While there are often advantages to an in-vivo system, organotypic slice cultures allow one to perform a number of manipulations that are not presently feasible in-vivo. To date, organtotypic embryonic brain slice cultures have been used to follow individual cells using time-lapse microscopy, manipulate the expression of genes in the ganglionic emanances (a region that is hard to target by in-utero electroporation), as well as for pharmacological studies. In this video protocol we demonstrate how to make organotypic slice cultures from rat embryonic day 18 embryos. The protocol involves dissecting the embryos, embedding them on ice in low melt agarose, slicing the embedded brains on the vibratome, and finally plating the slices onto filters in culture dishes. This protocol is also applicable in its present form to making organotypic slice cultures from different embryonic ages for both rats and mice. 

E18 랫 두뇌의 Organotypic 슬라이스 문화

Organotypic slice cultures from embryonic rodent brains are widely used to study brain development. While there are often advantages to an in-vivo system, ...

연구

생물학

Whole-cell Recordings of Light Evoked Excitatory Synaptic Currents in the Retinal Slice

We use the whole-cell patch clamp technique to study the synaptic circuitry that underlies visual information processing in the retina. In this video, we will guide you through the process of performing whole-cell recordings of light evoked currents of individual cells in the retinal slice preparation. We use the aquatic tiger salamander as an animal model. We begin by describing the dissection of the eye and show how slices are mounted for electrophysiological recordings. Once the slice is placed in the recording chamber, we demonstrate how to perform whole-cell voltage clamp recordings. We then project visual stimuli onto the photoreceptors in the slice to elicit light-evoked current responses. During the recording we perfuse the slice with pharmacological agents, whereby an 8-channel perfusion system allows us to quickly switch between different agents.  The retinal slice preparation is widely used for patch clamp recordings in the retina, in particular to study amacrine or bipolar cells, which are not accessible in a whole-mount preparation.

This video shows the process of whole-cell voltage clamp recordings in the retinal slice of the aquatic tiger salamander. We demonstrate the preparation of the slice as well as how to perform patch clamp recordings during visual stimulation of the retina.

라이트의 전체 세포 레코딩은 망막 통에 흥분성의 시냅틱 전류를 Evoked

We use the whole-cell patch clamp technique to study the synaptic circuitry that underlies visual information processing in the retina. In this video, we ...

Identifying Targets of Human microRNAs with the LightSwitch Luciferase Assay System using 3'UTR-reporter Constructs and a microRNA Mimic in Adherent Cells

MicroRNAs (miRNAs) are important regulators of gene expression and play a role in many biological processes. More than 700 human miRNAs have been identified so far with each having up to hundreds of unique target mRNAs. Computational tools, expression and proteomics assays, and chromatin-immunoprecipitation-based techniques provide important clues for identifying mRNAs that are direct targets of a particular miRNA. In addition, 3'UTR-reporter assays have become an important component of thorough miRNA target studies because they provide functional evidence for and quantitate the effects of specific miRNA-3'UTR interactions in a cell-based system. To enable more researchers to leverage 3'UTR-reporter assays and to support the scale-up of such assays to high-throughput levels, we have created a genome-wide collection of human 3'UTR luciferase reporters in the highly-optimized LightSwitch Luciferase Assay System. The system also includes synthetic miRNA target reporter constructs for use as positive controls, various endogenous 3'UTR reporter constructs, and a series of standardized experimental protocols.
Here we describe a method for co-transfection of individual 3'UTR-reporter constructs along with a miRNA mimic that is efficient, reproducible, and amenable to high-throughput analysis.

Identifying Targets of Human microRNAs with the LightSwitch Luciferase Assay System using 3&#039;UTR-reporter Constructs and a microRNA Mimic in Adherent Cells

MicroRNAs (miRNAs) are important regulators of gene expression and have been shown to play a role in numerous biological processes. To better understand miRNA-UTR interactions, we have created a genome-wide collection of 3 UTR luciferase reporters paired with a novel luciferase gene and assay reagent, the LightSwitch system.

3&#39;UTR - 기자의 구성과 microRNA를 사용하여 LightSwitch의 루시페라제 분석 시스템과 인간 microRNAs의 확인 대상이 자기편 세포의 모방

MicroRNAs (miRNAs) are important regulators of gene expression and play a role in many biological processes.  More than 700 human miRNAs have been ...

A Method for Selecting Structure-switching Aptamers Applied to a Colorimetric Gold Nanoparticle Assay

Small molecules provide rich targets for biosensing applications due to their physiological implications as biomarkers of various aspects of human health and performance. Nucleic acid aptamers have been increasingly applied as recognition elements on biosensor platforms, but selecting aptamers toward small molecule targets requires special design considerations. This work describes modification and critical steps of a method designed to select structure-switching aptamers to small molecule targets. Binding sequences from a DNA library hybridized to complementary DNA capture probes on magnetic beads are separated from nonbinders via a target-induced change in conformation. This method is advantageous because sequences binding the support matrix (beads) will not be further amplified, and it does not require immobilization of the target molecule. However, the melting temperature of the capture probe and library is kept at or slightly above RT, such that sequences that dehybridize based on thermodynamics will also be present in the supernatant solution. This effectively limits the partitioning efficiency (ability to separate target binding sequences from nonbinders), and therefore many selection rounds will be required to remove background sequences. The reported method differs from previous structure-switching aptamer selections due to implementation of negative selection steps, simplified enrichment monitoring, and extension of the length of the capture probe following selection enrichment to provide enhanced stringency. The selected structure-switching aptamers are advantageous in a gold nanoparticle assay platform that reports the presence of a target molecule by the conformational change of the aptamer. The gold nanoparticle assay was applied because it provides a simple, rapid colorimetric readout that is beneficial in a clinical or deployed environment. Design and optimization considerations are presented for the assay as proof-of-principle work in buffer to provide a foundation for further extension of the work toward small molecule biosensing in physiological fluids.

A protocol is provided to select structure-switching aptamers for small molecule targets based on a tunable stringency magnetic bead selection method. Aptamers selected with structure-switching properties are beneficial for assays that require a conformational change to signal the presence of a target, such as the described gold nanoparticle assay.

색채 금 나노 입자 분석에 적용 구조 전환 앱 타머를 선택하는 방법

Small molecules provide rich targets for biosensing applications due to their physiological implications as biomarkers of various aspects of human health ...

qPCRTag Analysis - A High Throughput, Real Time PCR Assay for Sc2.0 Genotyping

The Synthetic Yeast Genome Project (Sc2.0) aims to build 16 designer yeast chromosomes and combine them into a single yeast cell. To date one synthetic chromosome, synIII1, and one synthetic chromosome arm, synIXR2, have been constructed and their in vivo function validated in the absence of the corresponding wild type chromosomes. An important design feature of Sc2.0 chromosomes is the introduction of PCRTags, which are short, re-coded sequences within open reading frames (ORFs) that enable differentiation of synthetic chromosomes from their wild type counterparts. PCRTag primers anneal selectively to either synthetic or wild type chromosomes and the presence/absence of each type of DNA can be tested using a simple PCR assay. The standard readout of the PCRTag assay is to assess presence/absence of amplicons by agarose gel electrophoresis. However, with an average PCRTag amplicon density of one per 1.5 kb and a genome size of ~12 Mb, the completed Sc2.0 genome will encode roughly 8,000 PCRTags. To improve throughput, we have developed a real time PCR-based detection assay for PCRTag genotyping that we call qPCRTag analysis. The workflow specifies 500 nl reactions in a 1,536 multiwell plate, allowing us to test up to 768 PCRTags with both synthetic and wild type primer pairs in a single experiment.

Designer chromosomes of the Synthetic Yeast Genome project, Sc2.0, can be distinguished from their native counterparts using a PCR-based genotyping assay called PCRTagging, which has a presence/absence endpoint. Here we describe a high-throughput real time PCR detection method for PCRTag genotyping.

qPCRTag 분석 - 높은 처리량, Sc2.0 유전형에 대한 실시간 PCR 분석

The Synthetic Yeast Genome Project (Sc2.0) aims to build 16 designer yeast chromosomes and combine them into a single yeast cell. To date one synthetic ...

Design and Development of Aptamer&#8211;Gold Nanoparticle Based Colorimetric Assays for In-the-field Applications

The design and development of an aptamer&#8211;gold nanoparticle (AuNP) colorimetric assay for the detection of small molecules for in-the-field applications was examined. Target selective AuNP based color assays have been developed in controlled proof-of-concept laboratory settings. However, these schemes have not been exerted to a point of failure to determine their practical use beyond laboratory settings. This work describes a generic approach to design, develop, and troubleshoot an aptamer-AuNP colorimetric assay for small molecule analytes and using the assay for in-the-field settings. The assay is advantageous because adsorbed aptamers passivate the nanoparticle surfaces and provide a means to reduce and eliminate false positive responses to non-target analytes. Transitioning this system to practical uses required defining not only the shelf-life of the aptamer-AuNP assay, but establishing methods and procedures for extending the long-term storage capabilities. Also, one of the recognized concerns with colorimetric readout is the burden placed on analysts to accurately identify often subtle changes in color. To lessen the responsibility on analysts in the field, a color analysis protocol was designed to perform the color identification duties without the need for performing this task on laboratory grade equipment. The method for creating and testing the data analysis protocol is described. However to understand and influence the design of adsorbed aptamer assays, the interactions associated with the aptamer, target, and AuNPs require further study. The knowledge gained could lead to tailoring aptamers for improved functionality.

The design and development of an aptamer&#8211;gold nanoparticle colorimetric assay for the detection of small molecules for in-the-field applications was examined. Additionally, a smart-device colorimetric application (app) was validated and long-term storage of the assay was established for use in the field.

인 - 더 - 필드 응용 프로그램에 대한 설계 및 앱 타머 - 금 나노 입자 기반 비색 분석법 개발

The design and development of an aptamer&#8211;gold nanoparticle (AuNP) colorimetric assay for the detection of small molecules for in-the-field ...

The Assembly and Application of 'Shear Rings': A Novel Endothelial Model for Orbital, Unidirectional and Periodic Fluid Flow and Shear Stress

Deviations from normal levels and patterns of vascular fluid shear play important roles in vascular physiology and pathophysiology by inducing adaptive as well as pathological changes in endothelial phenotype and gene expression. In particular, maladaptive effects of periodic, unidirectional flow induced shear stress can trigger a variety of effects on several vascular cell types, particularly endothelial cells. While by now endothelial cells from diverse anatomic origins have been cultured, in-depth analyses of their responses to fluid shear have been hampered by the relative complexity of shear models (e.g., parallel plate flow chamber, cone and plate flow model). While these all represent excellent approaches, such models are technically complicated and suffer from drawbacks including relatively lengthy and complex setup time, low surface areas, requirements for pumps and pressurization often requiring sealants and gaskets, creating challenges to both maintenance of sterility and an inability to run multiple experiments. However, if higher throughput models of flow and shear were available, greater progress on vascular endothelial shear responses, particularly periodic shear research at the molecular level, might be more rapidly advanced. Here, we describe the construction and use of shear rings: a novel, simple-to-assemble, and inexpensive tissue culture model with a relatively large surface area that easily allows for a high number of experimental replicates in unidirectional, periodic shear stress studies on endothelial cells.

The Assembly and Application of &#039;Shear Rings&#039;: A Novel Endothelial Model for Orbital, Unidirectional and Periodic Fluid Flow and Shear Stress

Different levels and patterns of fluid shear are known to modulate endothelial gene expression, phenotype and susceptibility to disease. We discuss the assembly and use of 'shear rings': a model that produces unidirectional, periodic shear stress patterns. Shear rings are simple to assemble, economical and can produce high cell yields.

총회와 &#39;전단 반지&#39;의 응용 프로그램 : 궤도, 단방향 및 정기 유체 흐름 및 전단 응력에 대한 소설 내피 모델

Deviations from normal levels and patterns of vascular fluid shear play important roles in vascular physiology and pathophysiology by inducing adaptive as ...

Detecting and Characterizing Protein Self-Assembly In Vivo by Flow Cytometry

Protein self-assembly governs protein function and compartmentalizes cellular processes in space and time. Current methods to study it suffer from low-sensitivity, indirect read-outs, limited throughput, and/or population-level rather than single-cell resolution. We designed a flow cytometry-based single methodology that addresses all of these limitations: Distributed Amphifluoric FRET or DAmFRET. DAmFRET detects and quantifies protein self-assemblies by sensitized emission FRET in vivo, enables deployment across model systems-from yeast to human cells-and achieves sensitive, single-cell, high-throughput read-outs irrespective of protein localization or solubility.

This article describes a FRET-based flow cytometry protocol to quantify protein self-assembly in both S. cerevisiae and HEK293T cells.

유세포분석으로 생체내 단백질 자가 조립 검출 및 특성화

Protein self-assembly governs protein function and compartmentalizes cellular processes in space and time. Current methods to study it suffer from ...

Automated Charting of the Visual Space of Housefly Compound Eyes

This paper describes the automatic measurement of the spatial organization of the visual axes of insect compound eyes, which consist of several thousands of visual units called ommatidia. Each ommatidium samples the optical information from a small solid angle, with an approximate Gaussian-distributed sensitivity (half-width on the order of 1&#730;) centered around a visual axis. Together, the ommatidia gather the visual information from a nearly panoramic field of view. The spatial distribution of the visual axes thus determines the eye's spatial resolution. Knowledge of the optical organization of a compound eye and its visual acuity is crucial for quantitative studies of neural processing of the visual information. Here we present an automated procedure for mapping a compound eye's visual axes, using an intrinsic, in vivo optical phenomenon, the pseudopupil, and the pupil mechanism of the photoreceptor cells. We outline the optomechanical setup for scanning insect eyes and use experimental results obtained from a housefly, Musca domestica, to illustrate the steps in the measurement procedure.

The protocol here describes the measurement of the spatial organization of the visual axes of housefly eyes, mapped by an automatic device, using the pseudopupil phenomenon and the pupil mechanism of the photoreceptor cells.

Housefly Compound Eyes의 시각적 공간의 자동 차트 작성

This paper describes the automatic measurement of the spatial organization of the visual axes of insect compound eyes, which consist of several thousands ...

Collection and Long-Term Maintenance of Leaf-Cutting Ants (Atta) in Laboratory Conditions

Ants are one of the most biodiverse groups of animals on the planet and inhabit different environments. The maintenance of ant colonies in controlled environments enables an enriched comprehension of their biology that can contribute to applied research. This practice is usually employed in population control studies of species that cause economic loss, such as Atta ants. To cultivate their mutualistic fungus, these leaf-cutting ants collect leaves and for this are considered agricultural pests widely distributed throughout the American continent. They are highly socially organized and inhabit elaborated underground nests composed of a variety of chambers. Their maintenance in a controlled environment depends on a daily routine of several procedures and frequent care that are described here. It starts with the collection of queens during the reproductive season (i.e., nuptial flight), which are then individually transferred to plastic containers. Due to the high mortality rate of queens, a second collection can be carried out about 6 months after the nuptial flight, when incipient nests with developed fungus wad are excavated, hand-picked, and placed in plastic containers. In the laboratory, leaves are daily provided to established colonies, and ant-produced waste is weekly removed along with remaining dry plant material. As the fungus garden keeps growing, colonies are transferred to different types of containers according to the experimental purpose. Leaf-cutting ant colonies are placed in interconnected containers, representing the organizational system with functional chambers built by those insects in nature. This setup is ideal to monitor factors such as waste amount, fungus garden health, and the behavior of workers and queen. Facilitated data collection and more detailed observations are considered the greatest advantage of keeping ant colonies in controlled conditions.

Collection and Long-Term Maintenance of Leaf-Cutting Ants Atta in Laboratory Conditions

Here, a protocol is described to successfully collect and maintain healthy Atta (Hymenoptera: Formicidae) ant colonies in laboratory conditions. Additionally, different nest types and configurations are detailed together with possible experimental procedures.

실험실 조건에서 잎 절단 개미 (Atta)의 수집 및 장기 유지 관리

Ants are one of the most biodiverse groups of animals on the planet and inhabit different environments. The maintenance of ant colonies in controlled ...