J Cook

Overview:

I am interested in all aspects of flow cytometric analysis and cell sorting, especially the application of these techniques to elucidation of the development of the immune system and understanding primary immunodeficiencies. I am committed to assisting the Duke research community in the maximum utilization of the cytometers and techniques available to them through our shared reasource, including training of individuals to perform their own cytometry. My goal is to insure that their flow cytometry experiments will lead to peer reviewed publications and strong fundable RO1 applications.

Positions:

Assistant Research Professor of Immunology

Immunology
School of Medicine

Member of the Duke Cancer Institute

Duke Cancer Institute
School of Medicine

Education:

Ph.D. 1984

University of Virginia

Publications:

A comparison of immunohistochemistry, two-color immunofluorescence, and flow cytometry with cell sorting for the detection of micrometastatic breast cancer in the bone marrow.

A significant percentage of women with primary breast cancer have micrometastatic disease in the bone marrow. Bone marrow involvement may be an adverse prognostic factor, and more aggressive therapy may be indicated for these patients. There are a number of different techniques and antibodies used to detect tumor cells in the bone marrow. We used the same panels of four antibreast cancer antibodies and compared three immunodetection techniques: two-color immunofluorescence, immunohistochemical staining, and fluorescence-activated cell sorting with cytologic examination of the sorted cells. The two-color immunofluorescence technique was superior and consistently detected one tumor cell contaminating one million normal bone marrow cells and had no reactivity with normal bone marrow.
Authors
Vredenburgh, JJ; Silva, O; Tyer, C; DeSombre, K; Abou-Ghalia, A; Cook, M; Layfield, L; Peters, WP; Bast, RC
MLA Citation
Vredenburgh, J. J., et al. “A comparison of immunohistochemistry, two-color immunofluorescence, and flow cytometry with cell sorting for the detection of micrometastatic breast cancer in the bone marrow.J Hematother, vol. 5, no. 1, Feb. 1996, pp. 57–62. Pubmed, doi:10.1089/scd.1.1996.5.57.
URI
https://scholars.duke.edu/individual/pub749979
PMID
8646482
Source
pubmed
Published In
Journal of Hematotherapy
Volume
5
Published Date
Start Page
57
End Page
62
DOI
10.1089/scd.1.1996.5.57

Low temperature reversibly inhibits transport from tubular endosomes to a perinuclear, acidic compartment in African trypanosomes.

We have used electron microscopy and flow cytofluorimetry to study endocytosis and intracellular transport of fluid phase bovine serum albumen gold complexes and membrane bound concanavalin A through endosomal compartments of bloodstream forms of Trypanosoma brucei rhodesiense. Both markers were rapidly endocytosed from the flagellar pocket. Within 20 minutes at 37 degrees C the markers reached a large, vesicular, perinuclear compartment that stained heavily with the CB1 monoclonal antibody. Neither marker left the flagellar pocket and entered cells at 4 degrees C. When cells were incubated at 12 degrees C, both markers entered the cell and were transported to collecting tubules, a tubular endosomal compartment that receives endocytosed material from coated endocytic vesicles. However, no material was transported from collecting tubules to the late, perinuclear compartment at 12 degrees C. The morphology of collecting tubule membranes was specifically altered at 12 degrees C; tubules became shorter and were arrayed near the flagellar pocket. The morphological alteration and the block in transport of endocytic markers to the perinuclear compartment seen at 12 degrees C were reversed 10 minutes after cells were returned to 37 degrees C. We also used flow cytofluorimetric measurements of pH dependent fluorescence quenching to measure the pH of the terminal endocytic compartment. Fluoresceinated lectins accumulated in a terminal compartment with a pH of 6.0-6.1, a value considerably higher than that of mammalian lysosomes. Fluorescence from fluoresceinated lectins in this terminal endocytic compartment was dequenched when bloodstream forms were incubated in the presence of chloroquine.
Authors
Brickman, MJ; Cook, JM; Balber, AE
MLA Citation
Brickman, M. J., et al. “Low temperature reversibly inhibits transport from tubular endosomes to a perinuclear, acidic compartment in African trypanosomes.Journal of Cell Science, vol. 108 ( Pt 11), Nov. 1995, pp. 3611–21.
URI
https://scholars.duke.edu/individual/pub712700
PMID
8586672
Source
epmc
Published In
Journal of Cell Science
Volume
108 ( Pt 11)
Published Date
Start Page
3611
End Page
3621

A comparison of tumor and normal tissue microvascular hematocrits and red cell fluxes in a rat window chamber model.

This laboratory has previously used a window chamber model to measure red blood cell velocity in mammary tumors and normal granulation tissues of the F-344 rat. Because red cell flux and hematocrit more accurately reflect the oxygen carrying potential of blood, we used this model to measure these parameters. Red blood cells were labelled with fluorescein isothiocyanate, and 0.2 ml. packed cells were injected intravenously into rats bearing an 8 to 10 day old R-3230 mammary carcinoma. beta-phycoerythrin (0.15 mg.) was also injected and served as a plasma dye to outline the blood vessels. A sample of peripheral blood was then taken and analyzed by flow cytometry to determine the labeled fraction of red blood cells. Flowing tumor and normal tissue vessels were recorded onto a VCR, and these video images were used to determine vascular length and diameter, RBC flux and velocity, and hematocrit. Median vessel diameter and loge (red blood cell flux) were significantly greater in tumors than in normal tissues (p = 0.007 and p < 0.025, respectively). After controlling for these variables, the median tumor hematocrit of 19% was not significantly greater than the median normal tissue hematocrit of 15%. This technique provides a nontoxic and reproducible method that is now being used to assist in the in vivo definition of tumor oxygenation.
Authors
Brizel, DM; Klitzman, B; Cook, JM; Edwards, J; Rosner, G; Dewhirst, MW
MLA Citation
Brizel, D. M., et al. “A comparison of tumor and normal tissue microvascular hematocrits and red cell fluxes in a rat window chamber model.Int J Radiat Oncol Biol Phys, vol. 25, no. 2, Jan. 1993, pp. 269–76. Pubmed, doi:10.1016/0360-3016(93)90348-y.
URI
https://scholars.duke.edu/individual/pub712701
PMID
8420874
Source
pubmed
Published In
International Journal of Radiation Oncology, Biology, Physics
Volume
25
Published Date
Start Page
269
End Page
276
DOI
10.1016/0360-3016(93)90348-y

The beta-phosphoro[35S]thioate analogue of UDP-Glc is efficiently utilized by the glucose phosphotransferase and is relatively resistant to hydrolytic degradation.

The beta-phosphoro[35S]thioate analogue of UDP-glucose ((beta-35S)UDP-Glc) is utilized with approximately the same efficiency as the parent compound by the UDP-glucose:glycoprotein glucose-1-phosphotransferase (glucosyltransferase), which catalyzes the transfer of alpha Glc-1-P from UDP-Glc to mannose-containing oligosaccharides on acceptor glycoproteins. The same endogenous acceptor glycoproteins are labeled by the glucosyltransferase using [beta-32P]UDP-Glc and (beta-35S)UDP-Glc. However, in liver homogenates, incorporation from [beta-32P]UDP-Glc ceases to increase after about 4 min of incubation, while incorporation from (beta-35S)UDP-Glc persists for at least 1 h. This difference is due to an approx. 10-fold slower hydrolytic rate for the phosphorothioate analogue than for the parent compound, a finding similar to previous work showing that a variety of nucleases and phosphodiesterases are less efficient in cleaving phosphorothioate DNA than the native polymer.
Authors
Marchase, RB; Saunders, AM; Rivera, AA; Cook, JM
MLA Citation
Marchase, R. B., et al. “The beta-phosphoro[35S]thioate analogue of UDP-Glc is efficiently utilized by the glucose phosphotransferase and is relatively resistant to hydrolytic degradation.Biochim Biophys Acta, vol. 916, no. 2, Nov. 1987, pp. 157–62. Pubmed, doi:10.1016/0167-4838(87)90103-8.
URI
https://scholars.duke.edu/individual/pub712704
PMID
2823902
Source
pubmed
Published In
Biochimica Et Biophysica Acta
Volume
916
Published Date
Start Page
157
End Page
162
DOI
10.1016/0167-4838(87)90103-8

Amino acids stimulate cholecystokinin release through the Ca2+-sensing receptor.

Cholecystokinin (CCK) is produced by discrete endocrine cells in the proximal small intestine and is released following the ingestion of food. CCK is the primary hormone responsible for gallbladder contraction and has potent effects on pancreatic secretion, gastric emptying, and satiety. In addition to fats, digested proteins and aromatic amino acids are major stimulants of CCK release. However, the cellular mechanism by which amino acids affect CCK secretion is unknown. The Ca(2+)-sensing receptor (CaSR) that was originally identified on parathyroid cells is not only sensitive to extracellular Ca(2+) but is activated by extracellular aromatic amino acids. It has been postulated that this receptor may be involved in gastrointestinal hormone secretion. Using transgenic mice expressing a CCK promoter driven/enhanced green fluorescent protein (GFP) transgene, we have been able to identify and purify viable intestinal CCK cells. Intestinal mucosal CCK cells were enriched >200-fold by fluorescence-activated cell sorting. These cells were then used for real-time PCR identification of CaSR. Immunohistochemical staining with an antibody specific for CaSR confirmed colocalization of CaSR to CCK cells. In isolated CCK cells loaded with a Ca(2+)-sensitive dye, the amino acids phenylalanine and tryptophan, but not nonaromatic amino acids, caused an increase in intracellular Ca(2+) ([Ca(2+)](i)). The increase in [Ca(2+)](i) was blocked by the CaSR inhibitor Calhex 231. Phenylalanine and tryptophan stimulated CCK release from intestinal CCK cells, and this stimulation was also blocked by CaSR inhibition. Electrophysiological recordings from isolated CCK-GFP cells revealed these cells to possess a predominant outwardly rectifying potassium current. Administration of phenylalanine inhibited basal K(+) channel activity and caused CCK cell depolarization, consistent with changes necessary for hormone secretion. These findings indicate that amino acids have a direct effect on CCK cells to stimulate CCK release by activating CaSR and suggest that CaSR is the physiological mechanism through which amino acids regulate CCK secretion.
Authors
Wang, Y; Chandra, R; Samsa, LA; Gooch, B; Fee, BE; Cook, JM; Vigna, SR; Grant, AO; Liddle, RA
MLA Citation
Wang, Yu, et al. “Amino acids stimulate cholecystokinin release through the Ca2+-sensing receptor.Am J Physiol Gastrointest Liver Physiol, vol. 300, no. 4, Apr. 2011, pp. G528–37. Pubmed, doi:10.1152/ajpgi.00387.2010.
URI
https://scholars.duke.edu/individual/pub774327
PMID
21183662
Source
pubmed
Published In
Am J Physiol Gastrointest Liver Physiol
Volume
300
Published Date
Start Page
G528
End Page
G537
DOI
10.1152/ajpgi.00387.2010