Easy Question....5 points

T-cells express what primary CD marker(s)??

CD3 and CD4 or CD8

Source: CSM lecture notes from CCHMC

Easy Question.....5 points

True or False:

 A flow cytometer is the only way to look at surface antigen expression??

False.

There are several ways to look at surface expression of an antigen. One example would be using a fluorescent microscope.

Source: CSM lecture notes from CCHMC

Moderately hard question....10 points

The Specificity of an immunoassay is determined by the ..........

A. label used on the antigen

B. antibody used in the assay

C. Concentration of the unlabeled antigen

D. Concentration of the antibody

"B"

Specificity of an immunoassay is determined by the antibody used in the assay.

Specificity refers to the ability of an individual antibody combining site to react with only one antigenic determinant or ability of a population of antibody molecules to react with only one antigen.

Source: BOR Study Guide: ASCP

Easy question.....5 points

The most common label (fluorescent conjugate) used in direct fluorescent antibody techniques currently is....

A. PE

B. FITC

C. PerCP

D. PerCP cy5.5

"B"

FITC is the most common label used in direct fluorescent antibody techniques currently.

There are still many organizations that are using 4 color or less labeling techniques and therefore, FITC is the primary conjugate being used with  PE, APC, and PerCP.

Source: BOR Study Guide: ASCP

Easy question.....5 points

What is the proper name for FITC?

Fluorescein isothiocyanate

Source: SOP 7210-DLP-039-300

Easy question....5 points

An antibody is first added to the patient cells and incubated for the required amount of time. The cells are then washed. A fluorchrome is then added to the cells and incubated again for the required time period. This type of staining technique is called.....

A. direct fluorescent staining

B. indirect fluorescent staining

C. fluorescence inhibition staining

D. fluorescent quenching staining

"B"

Indirect fluoresecent staining

Indirect means the antibody has not been pre-conjugated with a fluorochrome. In order for the antibody to be detected a fluorochrome must be attached.

Source: BOR Study Guide: ASCP 

Hard Question....25 points

Avidity is defined as......

A. degreee of hemolysis

B. titer of an antigen

C. titer of an antibody

D. strength of a reacting antibody

"D"

Avidity is defined as the strength of a reacting antibody.

Another way to phrase this is....

Avidity is the interaction of an antibody with an antigen.

Source: BOR Study Guide: ASCP

Moderately Easy question....10 points

You run the Z-partical counter and get a WBC that is off the charts. You decide to dilute the sample 1:80 but upon recount you find the WBC is still too high. You take your first dilution and dilute that product again by 1:80. What is the number you should multiply your final count by to get the true WBC of this product?

(For those who need the technical issues: you take 10 ul and put it into 790 ul diluent, then take 10 ul of this product and put it into another vial of 790 ul diluent. This will give you two dilutions of 1:80)

6400

80 x 80 = 6400

If you were able to get an accurate count after the first 1:80 dilution you could have stopped and multiplied the WBC by 80. But because you had to make a second dilution of the product, you now need to multiple by an additional 80.

Source: CSM lecture notes from CCHMC

Moderately Easy Question....10 points

You have a WBC of 15.0 but to get this count you had to dilute the sample. You took 20 ul and placed it into 380 ul of diluent. What is your final WBC??

A. 150

B. 15.0

C. 300

D.5700

"C"

You took 20 ul of blood and added that to 380 ul diluent. This gives you a 1:20 dilution of the sample.

Then you would multiple the WBC of 15.0  by 20 to get a final WBC of 300.

Source: CSM lecture notes from CCHMC

Moderately Easy Question....10 points

You have a WBC that is very high so you take 5 ul and add this to 495 ul of diluent to bring the WBC down within an acceptable and readable range for the machine. What is your dilution factor?

1:100

Total volume = 500 ul

(5 ul of sample and 495 ul diluent)

500/5 =100

or another way to look at this

5 ul sample: 500 ul total dilution

Want to know for 1 ul of sample what amount of total diluent there is so divide both sides by "5"

Source: CSM lecture notes from CCHMC

TOUGH QUESTION!!! 25 points

Define hatenic determinants

 Hatenic determinants are subtances that are antigenic only when coupled to a protein carrier

Source: BOR Study Guide: ASCP

TOUGH QUESTION!!! 25 points!!

A hatenic determinant (a subtance that is antigenic only when coupled to a protein carrier) ...

A. Binds to B-cells while the carrier protion binds to the T-cells and B-cells are stimulated to produce hapten specific antibody

 OR

• B. Binds to T-cells while the carrier protion binds to B-cells and T-cells are stimulated to produce hapten specific antibody

"A"

A hatenic dterminant binds to the B-cells while the carrier portion binds to the T cells and B-cells are stimulated to produce hapten specific antibody.

Key to this question is: T-cells DO NOT produce the antibodies, B-cells do

Source: BOR Study Guide: ASCP

Easy Question....5 points

Cells that are precoursors of plasma cells and also produce immunoglobulinsa are....

A. Macrophages

B. B lymphocytes

C. T lymphocytes

D. Monocytes

"B"

B lymphocytes are precursors of plasma cells. B cells make antibodies, produce immunoglobulins, and then mature to plasma cells.

Source: BOR Study Guide: ASCP

Hard Question.....20 points

Immunoglobulin classes most commonly found on the surface of circulating B cells in peripheral blood are?

A. IgM and IgA

B. IgM and IgG

C. IgM and IgD

D. IgD and IgE

"C"

The most commonly found immunoglobulins found on the surface of circulating B cells in peripheral blood are IgM and IgD.

Source: BOR Study Guide: ASCP

Easy Question....5 points

CD4 cells are also called what type of cells?

Helper Cells

Source: BOR Study Guide: ASCP

Easy Question....5 points

CD8 cells are also called what type of cells?

Suppresor or cytotoxic T cells

Source: BOR Study Guide: ASCP

Moderately Easy Question...10 points

True or False:

CD3 cells do not have to commit to becoming CD4 or CD8 cells, they can just stay CD3+CD4-CD8- cells without any problems for the patient?

FALSE

CD3 must make a commitment to the CD4 pathway and become a helper cell or to the CD8 pathway and become a cytotoxic T cell. When a CD3 cell refuses to make the commitment one disease process that can occur is called ALPS (Acute lyphoproliferative syndrome).

Source: CSM Lecture notes from CCHMC

Moderately Easy Question...10 points

There is one type of lymphocyte that is self preservatory in nature meaning that if the bone marrow is shut down due to for example chemo, this cell type will find another organ within the body to hang out in until the marrow is functional again, using that organ as home to proliferate. What cell is this?

The T cell.

T cells are the only cells that must have a place within the body to proliferate. The cells may not be able to migrate to the thymus properly to make a commitment  but they will find some organ in the body to proliferate. Functionality is generally not there because the marrow has been shut down but the cells are attempting propigation some where to self preserve.

Source: CSM Lecture Notes from CCHMC

TOUGH QUESTION!! 25 points!!

Which cell is the "General" and coordinates most of the activities of the other cells?

The T Cell

T cells act as the General, telling other cells what they need to do and where they need to go within the body. If the other cells do not function properly, it is the T cell that will step in and try and take over the actions as can be seen for example in the NKT cells.

Source: CSM Lecture Notes from CCHMC

Easy Question...5 points

Yes or No

Do B cells need any stimulation or direction before the are funtional?

Yes

B-cells depend on the direction of the T-cells to perform their duties. B-cells are not cells that just jump and run to do a job. They wait for direction and leadership which comes from the T-cells.

Source: BOR Study Guide: ASCP

Easy Question.....5 points

True or False

NK cells need to recognize a bad cell before they will attack

FALSE

NK cells are like the basic private in an army. They are told to shoot and kill anything in site that appears to be a bad cell. The NK cells do not need to have been previous exposed to these bad cells to attack and destroy bad cells that are circulating. They only need to recognize the fact these are abnormal cells ( example: cells have been tagged previously by the spleen or liver marking for removal, or MHC class marker that is not of self).

Source: BOR Study Guide: ASCP

Easy Question....5 Points

True or False

T-cells can become phagocytic if need be

FALSE

This is probably the only function T cells cannot perform. T cells cannot be phagocytic and engulf debris and cells marked for destruction. However, T cells can become a natural killer (NKT) if necessary and perform a kill function. CD8 cells are also cytotoxic and perform specific kill functions.

Source: BOR Study Guide: ASCP

Easy Question.....5 points

True or False

By looking specifically at a CD3 marker you can identify which are helper cells and which are suppresor cells.

False

CD3 will only tell you if the cell is T cell or not. In order to know if it is a helper or suppresor cell, CD4 and or CD8 must be added to the cocktail.

If CD3 and CD4 are only added to the tube, most scientist assume the CD3+CD4- cell population is the CD3+CD8+ population. Remember tho if the doctor wants a quantitative number for CD8 and CD4 cells it is best to have CD4 and CD8 antibodies in the tube like we currently do here at Genoptix.  Using only CD3 and CD4 but determining CD8 presence through assumption is really a qualitative method not quanitative....and as you know there are always exceptions to everything!

Source: BOR Study Guide: ASCP

Hard Question.....

50 points if all answered correctly

5 points for each one correct

Note!!! Many answers have more than one answer

Match the follow:

A. Early T cell Marker                       

B. Mature T cell Marker                 

C. T cell Marker                    

D. Supporesor T cell Marker

E. Subsets of  T cells... Marker

F. Th cell Marker

G. NK cell Marker

H. Monocyte Marker

I.  B cell subset Marker

1. CD56

2. CD8

3. CD3

4. CD4

5. cyCD3

6. CD2

7. CD5

8. CD7

1. "E & G"

CD56- Found on subsets of T cells & NK cell marker

2. "D"

CD8- Suppressor or cytotoxic T cell marker

3. "B"

CD3- Mature T cell marker

4. "F & H"

CD4- Th (T- helper) cell and Monocyte marker

5. "A"

cyCD3- Early T cell marker

6. "A & G"

CD2- Early T cell and NK cell marker

7. "C & I"

CD5- T cell (norm & malignant) and B subset marker

8. "A & G"

CD7- Early T cell and NK cell marker

Source: CCHMC flow cytometry reports

Hard Question.....50 points if all correct

otherwise 5 points for each correct

Match the following:

A. Common ALL antigen marker

B. Early B cell marker

C. Ig Light Chain marker

D. Mature B cell marker

E. B cell marker

F. Plasma cell marker

1. CD79b

2. CD138

3. CD20

4. CD23

5. cyCD79a

6. CD22

7. CD19/Kappa or Lambda

8. CD19

9. CD10

1. "E"     CD79b- B cell marker

2. "F"     CD138- Plasma cell marker

3. "D"     CD20- Mature B cell marker

4. "D"     CD23- Mature B cell marker

5. "E"     cyCD79a- B cell marker

6. "E"     CD22- B cell marker

7. "C"     CD19/Kappa or Lambda- Ig light chains

8. "B"     CD19- Early B cell marker

9. "A"     CD10- common ALL antigen marker

Source: CCHMC Immunophentoyping report

Easy question....5 points

What does CD stand for?

Cluster designation or cluster of differentiaion.

Source http://www.pathologyoutlines.com/cdmarkers.html

Moderately hard Question....15 points

If you have an antibody and it is labeled for example:

wCD79a...what does the "w" mean?

When an antibody is classified, generally there are at least two monoclonal antibodies available for each antigen. The "w" means the cluster designation only has one monoclonal antibody or that the CD has not yet been well characterized and further research must be done.

http://www.pathologyoutlines.com/cdmarkers.html

Moderately Easy Question....10 points

Another name for CD2 and what type of marker is it?

E rosette receptor is another name for CD2. Also has the name LFA2 (leukocyte function antigen) and T11 (older names for this CD marker).

CD2 antibodies will inhibt the formation of rosettes when used with sheep erythrocytes.

CD2 is an Early T cell Marker

Source http://www.pathologyoutlines.com/cdmarkers.html

Tough Question....25 points for one correct answer. 15 points for each additionally correct answer

CD2 has 7 functions. Can you name one?

Functions of CD2 include:

(a) binds CD58 (LFA3) on antigen-presenting cells, which enables T cells to respond to lower concentrations of antigen (J Exp Med 1999;190:1383);

(b) induces costimulatory signals in T cells

(c) induces T cell cytokine production

(d) mediates adhesion between T cells and antigen presenting cells

(e) regulates T and NK-mediated cytolysis

(f) inhibits apoptosis of activated peripheral T cells

(g) regulates T cell anergy

SOURCE: http://www.pathologyoutlines.com/cdmarkers.html#CD2

Tough Question....25 points

Define Anergy...

"Anergy is a term in immunobiology that describes a lack of reaction by the body's defense mechanisms to foreign substances, and consists of a direct induction of peripheral lymphocyte tolerance. An individual in a state of anergy often indicates that the immune system is unable to mount a normal immune response against a specific antigen, usually a self-antigen. Lymphocytes are said to be anergic when they fail to respond to their specific antigen. Anergy is one of three processes that induce tolerance induction, modifying the immune system to prevent self-destruction (the others being clonal deletion and immunoregulation).^{[1]" WIKI}

http://en.wikipedia.org/wiki/Anergy

Easy question....5 points

Yes or No

Would you generally find postive CD2 staining on B cells, nonhematopoietic neoplasms or mast cells in non-mastocytosis disorders?

No: CD2 staining should be negative of B cells, nonhematopoietic neoplasms and mast cells in non-mastocytosis disorders.

http://www.pathologyoutlines.com/cdmarkers.html#CD2

Moderately hard question...15 points

What normal cells may pick up CD2?

"CD2 can be normally expressed on thymocytes (95%), mature peripheral T cells (almost all), NK cells (80-90%), and thymic B cells (50%)" (Wiki)

http://www.pathologyoutlines.com/cdmarkers.html#CD2

Moderately Easy Question...10 points

What is a thymocyte?

"Thymocytes are lymphocytes which develop in the thymus.^[1] They develop into T cells (but some evidence suggests they may also develop into natural killer cells.^{[2][3]" Wiki}).

http://en.wikipedia.org/wiki/Thymocytes

Moderately Easy Question....10 points

If a cell is a Thymocyte and it migrates to the thymus to develop (commit and mature), name CD marker combinations you could find on thymocytes.

Hint: talking about the general or most expected markers for thymocytes.

The following markers are the general ones found on Thymocytes

CD3+ CD4+ CD8-.....T helper cells

CD3+ CD4- CD8+.....T cytotoxic cells

CD3+ CD4- CD8-......Double neg T cells

CD3+ CD4+ CD8+....Double pos T cells

http://en.wikipedia.org/wiki/Thymocytes

Tough Question....25 points

Apoptosis for Thymocytes is an important process to keep those T cells that are non-functional out of circulation. Thymocytes go through ....

a.)thymus settling

b.) beta selection

c.) pos. selection & lineage commitment

d.) negative selection

Can you decribe in general tymus settling?

"The processes of beta-selection, positive selection and negative selection shape the population thymocytes into a peripheral pool of T cells that are able to respond to foreign pathogens and are immunologically tolerant towards self antigens" (Wiki).

a.) Thymus settling:

 "Thymocytes are dervived from the bone marrow but must reach the thymus to develop or commit to a cell lineage. To do so the thymus travels from the marrow cavity to the thymus through the circulation. The total number of progenitor cells that enter the thymus every day is estimated to be very small (don't need a lot of generals only a few good ones). It is at this point the progenitors proliferate to generate early T lineage cells. (CD3+CD4-CD8- cells)" (Wiki).

References:

http://en.wikipedia.org/wiki/Thymocytes

Tough Question....25 points

Apoptosis for Thymocytes is an important process to keep those T cells that are non-functional out of circulation. Thymocytes go through ....

a.)thymus settling

b.) beta selection

c.) pos. selection & lineage commitment

d.) negative selection

Can you decribe in general beta selection?

 "The processes of beta-selection, positive selection and negative selection shape the population thymocytes into a peripheral pool of T cells that are able to respond to foreign pathogens and are immunologically tolerant towards self antigens" (Wiki).

b.) Beta selection

"Beta selection eliminates cells with non-functional T cell receptors.The ability of T cells to recognize foreign antigens is mediated by the T cell receptor, which is a surface protein able to recognise short protein sequences (peptides) that are presented on MHC (major histocompatibility complex). During the double negative stage the major maturation step of thymocytes is to express a T cell receptor. The proteins encoded by the MHC are expressed on the surface of cells in all jawed vertebrates, and display both self antigens (peptide fragments from the cell itself) and nonself antigens (e.g., fragments of invading microorganisms) to a type of white blood cell called a T cell that has the capacity to kill or co-ordinate the killing of pathogens and infected or malfunctioning cells" (WIKI). 

References:

http://en.wikipedia.org/wiki/Major_histocompatibility_complex

http://en.wikipedia.org/wiki/Thymocytes

Tough Question....25 points

Apoptosis for Thymocytes is an important process to keep those T cells that are non-functional out of circulation. Thymocytes go through ....

a.)thymus settling

b.) beta selection

c.) pos. selection & lineage commitment

d.) negative selection

Can you decribe in general positive selection and lineage commitment?

"The processes of beta-selection, positive selection and negative selection shape the population thymocytes into a peripheral pool of T cells that are able to respond to foreign pathogens and are immunologically tolerant towards self antigens" (Wiki).

c.) pos. selection & lineage commitment

"The next major stage of thymocyte development is positive selection, to keep only those thymocytes which have a T cell receptor capable of binding MHC. The T cell receptor requires CD8 as a coreceptor to bind to MHC class I, and CD4 as a coreceptor to bind MHC class II. At this stage thymocytes upregulate both CD4 and CD8, becoming double positive cells.Thymocytes that have a T cell receptor incapable of binding MHC class I or class II undergo apoptosis. Some thymocytes are able to rescue failed positive selection by receptor editing (rearrangement of the other T cell receptor allele to produce a new T cell receptor)" (Wiki).

"The double positive thymocytes undergo lineage commitment, maturing into a CD8+ T cell (recognising MHC class I) or a CD4+ T cell (recognising MHC class II). Lineage commitment occurs at the late stage of positive selection and works by downregulation of both CD4 and CD8 (reducing the signal from the T cell receptor) and then upregulation of CD4 only. Thymocytes that start receiving signal again are those that recognise MHC class II, and they become CD4+ T cells. Thymocytes that do not start receiving signal again are those that recognize MHC class I, and they downregulate CD4 and upregulate CD8, to become CD8+ T cells. Both of these thymocytes types are known as single positive thymocytes" (Wiki).

http://en.wikipedia.org/wiki/Thymocytes

Tough Question....25 points

Apoptosis for Thymocytes is an important process to keep those T cells that are non-functional out of circulation. Thymocytes go through ....

a.)thymus settling

b.) beta selection

c.) pos. selection & lineage commitment

d.) negative selection

Can you decribe in general negative selection?

The processes of beta-selection, positive selection and negative selection shape the population thymocytes into a peripheral pool of T cells that are able to respond to foreign pathogens and are immunologically tolerant towards self antigens.

d.) negative selection

"At this stage the key maturation process involves negative selection, the elimination of autoreactive thymocytes. During negative selection, all thymocytes with a high affinity for binding self peptides presented on MHC class I or class II are induced to upregulate Bim, a protein which drives apoptosis. Cells which do not have a high affinity for self ('safe' cells) survive negative selection. Single positive thymocytes remain in the medulla for 1–2 weeks, surveying self-antigens to test for autoreactivity. During this time they undergo final maturational changes, and then exit the thymus using S1P and CCR7. Upon entry to the peripheral bloodstream, the cells are considered mature T cells, and not thymocytes." (wiki).

http://en.wikipedia.org/wiki/Thymocytes

Tough Question....25 points

What is an evolutionary advantage and disadvantage to having a large number of unique T cell receptors?

"The evolutionary advantage in having a large number of unique T cell receptors is that each T cell is capable of recognizing a different peptide, providing a defence against rapidly evolving pathogens.^{[7]" (Wiki)}.

"The cellular disadvantage in the rearrangement process is that many of the combinations of the T cell receptor gene fragments are non-functional. To eliminate thymocytes which have made a non-functional T cell receptor, the beta-selection point is required before T cells can advance from the double negative to the double positive stage. The beta-selection point requires that the first T cell receptor gene to be arranged (T cell receptor beta) is capable to binding a pre-T cell receptor alpha protein and assembling on the surface with the signalling proteins. Thymocytes which fail this "beta selection" die by apoptosis" (Wiki).

http://en.wikipedia.org/wiki/Thymocytes

Moderately hard question....20 points

A thymocyte has passed beta selection testing and has almost completed the positive selection and lineage commitment phase (is in the late stage of this step). There has been reduced signal from the T cell receptor followed by then an upregualation of one signal only.

A.) When the T cell receptor begins signally again... what cells will receive that signal?

B.) The cells that do not receive the signal, what happens to them?

 "Thymocytes that start receiving signal again are those that recognise MHC class II, and they become CD4+ T cells" (Wiki).

"Thymocytes that do not start receiving signal again are those that recognize MHC class I, and they downregulate CD4 and upregulate CD8, to become CD8+ T cells. Both of these thymocytes types are known as single positive thymocytes" (Wiki).

http://en.wikipedia.org/wiki/Thymocytes

Easy question.....5 points

At what wavelength is FITC excited and what wavelength does it emit?