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Combined flow cytometry and confocal laser scanning microscopy for evaluation of BR96 antibody cancer cell targeting and internalization.

Feridani, Amir LU ; Holmqvist, Bo LU ; Sjögren, Hans Olov LU ; Strand, Sven-Erik LU ; Tennvall, Jan LU and Baldetorp, Bo LU (2007) In Cytometry Part A 71A(6). p.361-370
Abstract
Background: Monoclonal antibodies (mAb) are important tools in the management of tumor disease, and the discovery of antibodies with both specific cancer cell targeting and capacity to enter the cells by internalization are critical to improve the therapeutic efficacy. Method: Antibody cancer cell targeting and internalization properties of fluoroscein-conjugated mAb made against Lewis Y (BR96) were evaluated quantitatively and qualitatively by means of flow cytometry (FCM) and confocal laser scanning microscopy (CLSM), respectively, on cells from a rat tumor cell line (BN7005-H1D2). Results: The study demonstrated a specific binding of BR96 to LewisY (LeY) located in the cell membrane and as BR96/LeY immunocomplexes (BR96/LeY)... (More)
Background: Monoclonal antibodies (mAb) are important tools in the management of tumor disease, and the discovery of antibodies with both specific cancer cell targeting and capacity to enter the cells by internalization are critical to improve the therapeutic efficacy. Method: Antibody cancer cell targeting and internalization properties of fluoroscein-conjugated mAb made against Lewis Y (BR96) were evaluated quantitatively and qualitatively by means of flow cytometry (FCM) and confocal laser scanning microscopy (CLSM), respectively, on cells from a rat tumor cell line (BN7005-H1D2). Results: The study demonstrated a specific binding of BR96 to LewisY (LeY) located in the cell membrane and as BR96/LeY immunocomplexes (BR96/LeY) internalized into the cytoplasm. BR96/LeY was internalized into about 15% of the cells, usually distributed throughout the cytoplasm, but also located close to the nuclei. Cytotoxic effects by BR96 were indicated, and CLSM visualized subpopulations containing cells with bound or internalized BR96/LeY that possessed morphologically pyknotic nuclei and disrupted DNA. Conclusion: The spatial-temporal pattern by BR96 cell targeting and internalization processes of BR96/LeY into the cancer cells expressing LeY was demonstrated by FCM and CLSM. Used together, the FCM and CLSM techniques provide a valuable tool for preclinical analyses of antibody targeting and their capacities as carriers of cytotoxic conjugates for the use in cancer therapy. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
confocal laser scanning microscopy, flow cytometry, Lewis Y antigen, immunoconjugate, cell death, antibody mediated internalization
in
Cytometry Part A
volume
71A
issue
6
pages
361 - 370
publisher
John Wiley & Sons
external identifiers
  • wos:000246718100003
  • scopus:34250736948
ISSN
1552-4930
DOI
10.1002/cyto.a.20388
language
English
LU publication?
yes
id
e3200561-d803-4b6b-aca4-a5e44d4cf945 (old id 166434)
alternative location
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=17366637&dopt=Abstract
date added to LUP
2007-07-13 13:48:37
date last changed
2017-01-01 04:51:56
@article{e3200561-d803-4b6b-aca4-a5e44d4cf945,
  abstract     = {Background: Monoclonal antibodies (mAb) are important tools in the management of tumor disease, and the discovery of antibodies with both specific cancer cell targeting and capacity to enter the cells by internalization are critical to improve the therapeutic efficacy. Method: Antibody cancer cell targeting and internalization properties of fluoroscein-conjugated mAb made against Lewis Y (BR96) were evaluated quantitatively and qualitatively by means of flow cytometry (FCM) and confocal laser scanning microscopy (CLSM), respectively, on cells from a rat tumor cell line (BN7005-H1D2). Results: The study demonstrated a specific binding of BR96 to LewisY (LeY) located in the cell membrane and as BR96/LeY immunocomplexes (BR96/LeY) internalized into the cytoplasm. BR96/LeY was internalized into about 15% of the cells, usually distributed throughout the cytoplasm, but also located close to the nuclei. Cytotoxic effects by BR96 were indicated, and CLSM visualized subpopulations containing cells with bound or internalized BR96/LeY that possessed morphologically pyknotic nuclei and disrupted DNA. Conclusion: The spatial-temporal pattern by BR96 cell targeting and internalization processes of BR96/LeY into the cancer cells expressing LeY was demonstrated by FCM and CLSM. Used together, the FCM and CLSM techniques provide a valuable tool for preclinical analyses of antibody targeting and their capacities as carriers of cytotoxic conjugates for the use in cancer therapy.},
  author       = {Feridani, Amir and Holmqvist, Bo and Sjögren, Hans Olov and Strand, Sven-Erik and Tennvall, Jan and Baldetorp, Bo},
  issn         = {1552-4930},
  keyword      = {confocal laser scanning microscopy,flow cytometry,Lewis Y antigen,immunoconjugate,cell death,antibody mediated internalization},
  language     = {eng},
  number       = {6},
  pages        = {361--370},
  publisher    = {John Wiley & Sons},
  series       = {Cytometry Part A},
  title        = {Combined flow cytometry and confocal laser scanning microscopy for evaluation of BR96 antibody cancer cell targeting and internalization.},
  url          = {http://dx.doi.org/10.1002/cyto.a.20388},
  volume       = {71A},
  year         = {2007},
}