Name: using mouse mammary tumor cells to teach core biology concepts: a simple lab module
Uploaded: 2015-06-18T13:00:00
Description: Watch this Scientific Journal Video about Using Mouse Mammary Tumor Cells to Teach Core Biology Concepts: A Simple Lab Module at JoVE.com

This work is supported by the Joseph Alexander Foundation, the ASBMB Undergraduate Research Award, 2013-2014, and a Science Award Grant, Marymount Manhattan College, 2012-2013.

Bemærkninger: Adfærd alt arbejde med celler og cellekulturreagenser i en klasse II biosikkerhed kabinet (BSC) 9. MMT celler klassificeres som biosikkerhed niveau I, da de udgør lav til moderat biologisk risiko. Påfør ordentlig rengøring og dekontaminering procedurer til BSC mellem anvendelser (f.eks ultraviolet lys, 70% ethanol tørre ned). 1. Grow MMT celler <ol><li> Grow celler i 10 cm vævskulturskåle indeholdende 10 ml næringsrige medier, der består af ...

<ol>
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En lab modul præsenteres der sigter mod at undervise i en række emner i cellebiologi gennem de avancerede teknikker af animalsk cellekultur. Modulet opnår dette ved at analysere virkningerne af en række anti-proliferative kemikalier på replikationen af ​​celler, der modellerer human brystcancer. Det primære assay er afhængig af den grundlæggende teknik med celletælling og indfører en ny måde at tælle celler under anvendelse mikroskopi software. Aktiviteterne omfatter modulet kan udføres med instrumenter...

Ofte i bachelor generel biologi kurser, er emner af cellecyklusregulering og kræft berørt, men ikke undersøgt i detaljer, fordi bredden af ​​indholdet i disse kurser giver lidt tid til dybde. Desuden er bachelorstuderende biologistuderende ikke typisk udsat for de avancerede teknikker er forbundet med animalsk cellekultur. For at hjælpe eleverne med at udvikle en dybere forståelse af disse begreber, mens anvendelse og analysere, hvad de har lært, blev et laboratorium, der udvikles som en modifikation af Walter Reed Army Institute of Research (WRAIR) udvidet laboratorieaktivitet 1. Laboratoriet modul anvender en trinvis, eksperimentel strategi, som omfatter dyrkning og karakterisering en cancercelle model, udvikle og gennemføre celletælling metoder, oprettelse optimal tidsforløb og doseringer til behandling af celler med anti-proliferative midler, og identificere aberrerende celle-signalveje . Eksperimentet giver også mulighed for åben-ended undersøgelse. De fleste af de teknikker, der kræves til denne aktivitet kan udføres i et typisk biologi-undervisning laboratorium. Aktiviteten starter med studerende karakteriserer morfologi og vækstrate for musebrysttumor (MMT) cellelinje, en model for human brystkræft 2. Brystkræft blev valgt som model kræft på grund af sin udbredelse i befolkningen, dens kendskab til college-alderen studerende, og de udbredte data. MMT cellelinjen blev specifikt valgt, fordi det er vanskeligt at indhente velkarakteriseret, har en kort fordoblingstid og er nemme at dyrke. Desuden MMT celler er østrogen-afhængige som er i overensstemmelse med de fleste kvindelige brystkræft. Studerende derefter identificere afvigende celle-signalveje i MMT celler ved behandling af cellerne med kemoterapi narkotika, hvis virkningsmekanisme er godt established.The koncentration af narkotika og længden af ​​behandlinger varierede giver eleverneat evaluere virkningen af ​​disse variable på hastigheden af ​​celledeling. Nøglen assay for denne aktivitet er bestemmelsen af ​​cellelevedygtighed, som blot kræver celletælling, ved hjælp af en af ​​to metoder. Hver metode afhænger af stærke mikroskopi færdigheder. Studerende bestemme cellelevedygtighed ved anvendelse af en standard, hæmocytometer fremgangsmåde og et hidtil ukendt fotomikroskopi fremgangsmåde og foreslår. Baseret på deres resultater, kan de foreslå og afprøve ændringer til aktiviteten. Studerende derefter repræsentere deres data og fortolke resultaterne til at forfine deres hypotese og udtænke nye eksperimentelle strategier. Dette laboratorium aktivitet er velegnet til freshman eller sophomore niveau studerende med hovedfag i de biologiske videnskaber. Den er kondenseret til en uges lab modul, der kan være afsluttet i en første år, generel biologi eller andet år, cellulær / molekylær biologi kursus. Færdigheder er nødvendige for korrekt gennemførelse af aktiviteten omfatter grundlæggende aritmetik og algebra, kendskab til en bred vifte af cmalm laboratorie færdigheder (f.eks pipettering, løsning gør, steril teknik), dataanalyse, grundlæggende lysmikroskopi og tidsstyring, sammen med instruktør kendskab til cellekultur og regneark. Reagenser nødvendige omfatter et dyr cellelinie model for cancer (f.eks muse brysttumorceller, MMT 2), kemoterapeutiske stoffer (f.eks, tamoxifen, curcumin, metformin, og aspirin), trypanblåt og celledyrkningsmedier (f.eks Eagles Minimum Essential Medium ; EMEM) med passende kosttilskud (f.eks donor hest og føtalt bovint serum). Nødvendige instrumenter omfatter en inverteret lysmikroskop med digital kamera vedhæftet fil, computer, 100 mm og 24 brønds vævskulturplader, CO2-inkubator (eller tilsvarende), biosikkerhed kabinet (BSC; klasse II), hæmocytometer, og digital mikroskopi software. Der er gode eksempler på specifikke lab aktiviteter, der er afhængige af dyrecellekultur at TEACh studerende om begreber i cellebiologi 3. Men mange kræver forsyninger eller teknikker, som ikke er let tilgængelige (fx radioaktive isotoper, levende dyr væv, avanceret billedbehandling udstyr 1,4,5), beskriver protokoller, der er ganske fremskreden (fx egnet til en 400-niveau kursus 6), eller kræver multi-ugers eller semester lange projekter 6,7. Laboratoriet aktivitet er beskrevet her, er ligetil og kan udføres i en enkelt uge med fælles lab udstyr. Sammenfattende denne lab modul effektivt introducerer eller styrker begreberne cellecyklus, cellulære signalveje og kræft mens undervisning grundlæggende og avancerede lab færdigheder, eksperimentel dataanalyse, metoden til dyrecellekultur og den videnskabelige proces. Laboratoriet modulet er enkel og økonomisk tilgængelig og giver både fleksibilitet og mulighed for åben undersøgelse. Aktiviteten tilskynder studerende kreativitetved at tilvejebringe en skabelon eksperimentel strategi, der fungerer som en vejledning, men ikke en opskrift. Vigtigst, aktivitet opfylder alle læringsområder af Blooms taksonomi 8, da det kræver at huske, forståelse, anvendelse, analyse og evaluering og skabe ved at engagere de studerende i en proces, der trækker dem ud af lærebogen og ind i verden af videnskabelig forskning.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Tissue Culture Hood</td>
			<td>ESCO Labculture Reliant</td>
			<td></td>
			<td>Class II Type A2 Biological Safety Cabinet</td>
		</tr>
		<tr>
			<td>Waterjactor CO2 Incubator</td>
			<td>CEDCO</td>
			<td>Model 1510</td>
			<td></td>
		</tr>
		<tr>
			<td>Bright-line Hemocytometer</td>
			<td>American Optical</td>
			<td></td>
			<td>with two separate grids</td>
		</tr>
		<tr>
			<td>Motic Images Plus</td>
			<td></td>
			<td></td>
			<td>Mac OSX Verison 2.0 or higher</td>
		</tr>
		<tr>
			<td>Gilson Pipetman</td>
			<td>Rainin instrument co. inc</td>
			<td></td>
			<td>P-20D, P-200D, P-1000D</td>
		</tr>
		<tr>
			<td>CK30/CK40 Culture Microscope</td>
			<td>Olympus</td>
			<td></td>
			<td>4 objective inverted light microscope with camera</td>
		</tr>
		<tr>
			<td>200 uL Pipet tips</td>
			<td>MidSci</td>
			<td>40200C</td>
			<td></td>
		</tr>
		<tr>
			<td>1000 uL Pipet tips</td>
			<td>MidSci</td>
			<td>AVR4</td>
			<td></td>
		</tr>
		<tr>
			<td>10 mL Seriological Pipets</td>
			<td>TPP</td>
			<td>TP94010</td>
			<td></td>
		</tr>
		<tr>
			<td>24-well plates</td>
			<td>CoStar- Tissue Culture Cluster</td>
			<td align="right">3524</td>
			<td>24 wells, 16 mm well diameter, Radiation sterilized</td>
		</tr>
		<tr>
			<td>Trypan Blue Solution 0.4%</td>
			<td>Sigma</td>
			<td>T8154</td>
			<td>100 mL, cell culture tested non-haz</td>
		</tr>
		<tr>
			<td>Bright-line Hemacytometer replacement coverslip, non-haz</td>
			<td>Sigma</td>
			<td>Z375357</td>
			<td></td>
		</tr>
		<tr>
			<td>Mouse Mammary Tumor(MMT) cells</td>
			<td>ATCC</td>
			<td>CCL-51</td>
			<td></td>
		</tr>
		<tr>
			<td>Eagle Minimum Essentail Medium (EMEM)</td>
			<td>ATCC</td>
			<td>30-2003</td>
			<td>500 mL</td>
		</tr>
		<tr>
			<td>Fetal Bovine Serum</td>
			<td>Sigma</td>
			<td>F0926</td>
			<td>500 mL</td>
		</tr>
		<tr>
			<td>Meformin Hydrochloride</td>
			<td>Sigma</td>
			<td>PHR1084</td>
			<td>500 mg</td>
		</tr>
		<tr>
			<td>Tamoxifen</td>
			<td>Sigma</td>
			<td>T5648</td>
			<td>white or white-yellow powder</td>
		</tr>
		<tr>
			<td>Curmumin</td>
			<td>Sigma</td>
			<td>C1386</td>
			<td>yellow-orange powder</td>
		</tr>
		<tr>
			<td>Aspirin</td>
			<td>Sigma</td>
			<td>A2093</td>
			<td>meets USP testing specifications</td>
		</tr>
	</tbody>
</table>

using mouse mammary tumor cells to teach core biology concepts: a simple lab module

Undergraduate biology students are required to learn, understand and apply a variety of cellular and molecular biology concepts and techniques in preparation for biomedical, graduate and professional programs or careers in science. To address this, a simple laboratory module was devised to teach the concepts of cell division, cellular communication and cancer through the application of animal cell culture techniques. Here the mouse mammary tumor (MMT) cell line is used to model for breast cancer. Students learn to grow and characterize these animal cells in culture and test the effects of traditional and non-traditional chemotherapy agents on cell proliferation. Specifically, students determine the optimal cell concentration for plating and growing cells, learn how to prepare and dilute drug solutions, identify the best dosage and treatment time course of the antiproliferative agents, and ascertain the rate of cell death in response to various treatments. The module employs both a standard cell counting technique using a hemocytometer and a novel cell counting method using microscopy software. The experimental procedure lends to open-ended inquiry as students can modify critical steps of the protocol, including testing homeopathic agents and over-the-counter drugs. In short, this lab module requires students to use the scientific process to apply their knowledge of the cell cycle, cellular signaling pathways, cancer and modes of treatment, all while developing an array of laboratory skills including cell culture and analysis of experimental data not routinely taught in the undergraduate classroom.

Voksende MMT celler og sammenligne tællemetoder. Musebrysttumor celler lykkedes dyrket og karakteriseret (figur 1) og en hidtil ukendt celletælling metode udviklet under anvendelse Motic Software, et digitalt kamera-associeret softwareprogram til et mikroskop. Denne nye celletælling metode blev sammenlignet med en traditionel optælling metode anvender et hæmocytometer (figur 2) og blev vist at være lige nøjagtig ved bestemmelse celletal <stro...

Watch this Scientific Journal Video about Using Mouse Mammary Tumor Cells to Teach Core Biology Concepts: A Simple Lab Module at JoVE.com

Using Mouse Mammary Tumor Cells to Teach Core Biology Concepts: A Simple Lab Module

A feasible laboratory module for biology undergraduates that explores advanced cellular and molecular concepts using animal cell culture is described. Students grow, characterize and manipulate a breast cancer cell model by exposure to chemotherapy agents. Cell viability is assayed through cell counting using both a standard and novel method.

Ved hjælp af musen brysttumorceller at undervise Core Biologi Begreber: A Simple Lab Modul

Undergraduate biology students are required to learn, understand and apply a variety of cellular and molecular biology concepts and techniques in ...

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