1887

Using Flow Cytometry to Measure Phagocytic Uptake in Earthworms

    Author: Sheryl L. Fuller-Espie1
    VIEW AFFILIATIONS HIDE AFFILIATIONS
    Affiliations: 1: Science Department, Cabrini College, Radnor, PA 19087
    AUTHOR AND ARTICLE INFORMATION AUTHOR AND ARTICLE INFORMATION
    • Published 20 December 2010
    • Supplemental material available at http://jmbe.asm.org
    • *Corresponding author. Mailing address: Science Department, Cabrini College, 610 King of Prussia Road, Radnor, PA 19087-3698. Phone: (610) 902-8369. Fax: (610) 902-8285. E-mail: [email protected].
    • Copyright © 2010 American Society for Microbiology
    Source: J. Microbiol. Biol. Educ. December 2010 vol. 11 no. 2 144-151. doi:10.1128/jmbe.v11i2.135
MyBook is a cheap paperback edition of the original book and will be sold at uniform, low price.
  • XML
  • HTML
    55.87 Kb
  • PDF
    237.22 Kb

    Abstract:

    This laboratory module familiarizes students with flow cytometry while acquiring quantitative reasoning skills during data analysis. Leukocytes, also known as coelomocytes (including hyaline and granular amoebocytes, and chloragocytes), from (earthworms) are isolated from the coelomic cavity and used for phagocytosis of fluorescent . Students learn how to set up cellular assays and become familiar with theoretical principles of flow cytometry. Histograms based on fluorescence and scatter properties combined with gating options permit students to restrict their analyses to particular subsets of coelomocytes when measuring phagocytosis, a fundamentally important innate immune mechanism used in earthworms. Statistical analysis of data is included in laboratory reports which serve as the primary assessment instrument.

Key Concept Ranking

Innate Immune System
0.471044
Polycyclic Aromatic Hydrocarbons
0.44493854
Chemicals
0.44211417
Reactive Oxygen Species
0.427141
Immune Response
0.42332727
0.471044

References & Citations

1. Axline SG, Reaven EP 1974 Inhibition of phagocytosis and plasma membrane mobility of the cultivated macrophage by Cytochalasin B: role of subplasmalemmal microfilaments J Cell Biol 62 647 659 10.1083/jcb.62.3.647 4368822 http://dx.doi.org/10.1083/jcb.62.3.647
2. Cooper EL, Kauschke E, Cossarizza A 2002 Digging for innate immunity since Darwin and Metchnikoff BioEssays 24 319 333 10.1002/bies.10077 11948618 http://dx.doi.org/10.1002/bies.10077
3. Engelmann P, Molnár L, Pálinkás L, Cooper EL, Németh P 2004 Earthworm leukocyte populations specifically harbor lysosomal enzymes that may respond to bacterial challenge Cell and Tissue Res 316 391 401 10.1007/s00441-004-0874-x http://dx.doi.org/10.1007/s00441-004-0874-x
4. Forman HJ, Torres M 2002 Reactive oxygen species and cell signaling: respiratory burst in macrophage signaling Am J Respir Crit Care Med 166 S4 S8 10.1164/rccm.2206007 12471082 http://dx.doi.org/10.1164/rccm.2206007
5. Fuller-Espie SL, Goodfield L, Hill K, Grant K, DeRogatis N 2008 Conservation of cytokine-mediated responses in innate immunity: a flow cytometric study investigating the effects of human proinflammatory cytokines on phagocytosis in the earthworm Eisenia hortensis Invertebrate Survival Journal 5 124 134 [Online] http://www.isj.unimore.it/
6. Givan AL 2001 Flow cytometry, first principles 2nd ed 15 57 John Wiley & Sons, Inc. New York 10.1002/0471223948.ch3 http://dx.doi.org/10.1002/0471223948.ch3
7. Jarosz J, Glinski Z 1997 Earthworm immune responses Folia Biologica 45 1 9
8. Malawista SE, Gee JB, Bensch KG 1971 Cytochalasin B reversibly inhibits phagocytosis: functional, metabolic, and ultrastructural effects in human blood leukocytes and rabbit alveolar macrophages Yale J Biol Med 44 286 300 5132788
9. Segal AW 2005 How neutrophils kill microbes Annu Rev Immunol 23 197 223 10.1146/annurev.immunol.23.021704.115653 15771570 http://dx.doi.org/10.1146/annurev.immunol.23.021704.115653
10. Underhill DM, Ozinsky A 2002 Phagocytosis of microbes: complexity in action Annu Rev Immunol 20 825 852 10.1146/annurev.immunol.20.103001.114744 11861619 http://dx.doi.org/10.1146/annurev.immunol.20.103001.114744

Supplemental Material

Loading

Article metrics loading...

/content/journal/jmbe/10.1128/jmbe.v11i2.135
2010-12-20
2019-02-21

Abstract:

This laboratory module familiarizes students with flow cytometry while acquiring quantitative reasoning skills during data analysis. Leukocytes, also known as coelomocytes (including hyaline and granular amoebocytes, and chloragocytes), from (earthworms) are isolated from the coelomic cavity and used for phagocytosis of fluorescent . Students learn how to set up cellular assays and become familiar with theoretical principles of flow cytometry. Histograms based on fluorescence and scatter properties combined with gating options permit students to restrict their analyses to particular subsets of coelomocytes when measuring phagocytosis, a fundamentally important innate immune mechanism used in earthworms. Statistical analysis of data is included in laboratory reports which serve as the primary assessment instrument.

Highlighted Text: Show | Hide
Loading full text...

Full text loading...

/deliver/fulltext/jmbe/11/2/jmbe-11-2-144.xml.a.html?itemId=/content/journal/jmbe/10.1128/jmbe.v11i2.135&mimeType=html&fmt=ahah

Figures

Image of FIGURE 1

Click to view

FIGURE 1

Shown are coelomocytes from after extrusion from the coelomic cavity. The large cells resembling fried eggs are the hyaline amoebocytes (large coelomocytes). The smallest cells with the white halo around their perimeter are the granular amoebocytes (small coelomocytes). The cells containing multiple small vesicular inclusions are the chlorogocytes (eleocytes). (Phase contrast microscopy, 400×)

Source: J. Microbiol. Biol. Educ. December 2010 vol. 11 no. 2 144-151. doi:10.1128/jmbe.v11i2.135
Download as Powerpoint
Image of FIGURE 2

Click to view

FIGURE 2

A forward (FSC) versus side (SSC) scatter profile of coelomocytes from . Note the rings [called regions (R)] drawn around the three major subpopulations: R1 = hyaline amoebocytes; R2 = granular amoebocytes; and R3 = chloragocytes. A region is a boundary drawn around a subpopulation for analysis. Once the regions have been established, analysis (e.g., FSC versus fluorescence detected by the FL-1 photomultiplier tube) can be carried out specifying which particular regions to include, a process known as gating. A gate may include one region or more than one region.

Source: J. Microbiol. Biol. Educ. December 2010 vol. 11 no. 2 144-151. doi:10.1128/jmbe.v11i2.135
Download as Powerpoint
Image of FIGURE 3

Click to view

FIGURE 3

Results of pre-test and post-test are illustrated. Left to right: pre-test for all students in class; post-test for all students in class; pre-test for students in class excluding students enrolled in BIO 312 (Theory & Practice in Biotechnology); post-test for students in class excluding students enrolled in BIO 312.

Source: J. Microbiol. Biol. Educ. December 2010 vol. 11 no. 2 144-151. doi:10.1128/jmbe.v11i2.135
Download as Powerpoint

This is a required field
Please enter a valid email address
Please check the format of the address you have entered.
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error