6 Chromosome Analysis

CHROMOSOME ANALYSIS

Used to look at the chromosomes in a sample of cells.

It can help identify the genetic abnormalities as the cause of a condition or disease

Karyotype

The chromosomal constitution of an individual.

It is also used to describe a photomicrograph of an individual's chromosomes arranged in a standard manner

Karyogram

Figure/diagram representing the karyotype of an individual

metaphase
vitro

To obtain ______ cells for chromosome analysis, patient somatic cells are cultured in____.

24
1

The average human cell divides once every __ hours, so only about _% of the cell population is dividing at any given time.

viable, nucleated

Virtually any ___, _____cell sample can be used for cytogenetic analysis

Heparinized peripheral blood

Preferred specimen for routine cytogenetic studies.

Bone marrow samples

Best results for hematologic disorders.

Fibroblasts from skin biopsies

Adequate source of metaphase cells.

Tissues such as liver

Not routinely used; however, these frequently provide an excellent resource if obtained soon after death during autopsy or from a fetal loss.

Amniotic fluid cells

Most common specimen for prenatal analysis.

Chorionic villi

Also for prenatal analysis

Percutaneous blood

Also for prenatal analysis; fetal blood specimen for rapid karyotyping or molecular studies.

sterile

All clinical samples for cytogenetic studies must be handled in a ____ manner

room temperature

Blood, bone marrow, amniotic fluid, and chorionic villi: Transported at

wet ice

Solid tissue: Transported on

Suspension (floating)

Culture technique depends on the cell type:
blood and bone marrow

Monolayer (fixed to a surface)

Culture technique depends on the cell type:
amniotic fluid cells, chorionic villi, and solid tissue

24–48 hours

Culture time per sample:
Bone marrow

3–4 days

Culture time per sample
Blood (lymphocytes)

5–7 days

Culture time per sample
Amniocytes and chorionic villi

Up to 2 weeks

Culture time per sample
Solid tissue culture

5

Procedure for suspension culture (blood/lymphocytes):

Collection: _ mL of heparinized venous blood is collected under sterile conditions

37

Procedure for suspension culture (blood/lymphocytes):

Incubation: Culture vial is placed in an incubator at __ °C.

mitotic inhibitor, colcemid

Procedure for suspension culture (blood/lymphocytes):

At the end of the prescribed culture time, a ____ ______, _______, is added into the culture vial.

Hypotonic treatment

Cells are treated with hypotonic saline, which causes cells to swell and chromosomes to separate.

glacial acetic acid and methanol

Procedure for suspension culture (blood/lymphocytes):

Fixation: Cells are fixed by adding a mixture of ____ ____ ____ and ____.

chilled

Procedure for suspension culture (blood/lymphocytes):

Slide preparation: Fixed cells are dropped on ____ slides from a specified height.

CHROMOSOME BANDING/STAINING

The chromosomes are visualized by the use of traditionally dyes, which stain them uniformly and provide uniform coloration to the chromosomes.

G-BANDING

Most common method; adequate for most situations.

Chromosomes are first treated with trypsin; slides are then stained with Giemsa solution, which stains each chromosome, showing a unique pattern of alternating light and dark regions/bands.

trypsin
Giemsa

G-BANDING

Chromosomes are first treated with ___; slides are then stained with ___ solution, which stains each chromosome, showing a unique pattern of alternating light and dark regions/bands.

Q-BANDING

uses quinacrine mustard stain and requires a UV fluorescent microscope. It produces a banding pattern similar to G-banding and is mainly used today for quick identification of the Y chromosome, especially in cases of ambiguous genitalia.

quinacrine mustard stain
UV fluorescent

Q-BANDING

Uses ___ ___ ___; was originally used in routine chromosome analysis.

Banding pattern is similar to G-banding, but slides have to be observed under a __ ___ microscope

Y

Q-banded preparation can usually determine the presence or absence of a _ chromosome in the patient.

Giemsa
reversed
deletions

R-BANDING
Uses ___ stain; however, chromosomes are pre-heated prior to staining.

Gives a banding pattern that is ___ of the G-banding.

Used in the detection of ___ not easily seen in G-banding.

R-BANDING

Uses Giemsa stain; however, chromosomes are pre-heated prior to staining.

Gives a banding pattern that is reversed of the G-banding.

Used in the detection of deletions not easily seen in G-banding.

C-BANDING

Regions of the centromere and secondary constrictions are stained.

Used to evaluate constitutive heterochromatin or to determine whether a chromosome has two centromeres (dicentric).

In the case of a dicentric chromosome, the presence of two dark regions clearly identifies the two centromeres.

centromere and secondary constrictions

C-BANDING
Regions of the ___ and ___ ___ are stained.

KARYOTYPE ANALYSIS

(cytogenetic) analysis, it is essential to rapidly and accurately identify each chromosome and determine when chromosome abnormalities are present.

active centromeres

The number of ___ ___ defines the total number of chromosomes, which will total 46 in a normal human diploid cell.

numerical abnormality

Too many or too few chromosomes indicate a potential ___ ___.

15–20

In vitro culture can result in culture artifacts, so no single cell is used to define an individual's chromosome complement. A typical clinical study requires analysis of __-__ cells.

mosaicism
10–30

In cases of ___ or other special situations, __-__  additional cells may be evaluated.

G

In most circumstances, routine _-banding of metaphase chromosomes is sufficient for clinical diagnostic purposes.

p arm

shorter arm
up

q arm

longer arm
down

computer-assisted imaging

Currently, most cytogenetics laboratories have moved to ___-___ ___

standard frame grabber

Using specially designed software, an image of a metaphase cell is captured using a ___ ___ ___, digitized, and displayed on the computer monitor

pattern-recognition subroutine

A karyogram can then be generated using a ___-___ ___, which will automatically identify the chromosomes and place them in their proper places on a karyogram form.