Bronchial Washing

 Purpose:To detect and classify neoplasms, involving the bronchial tree. Useful for
peripheral lung lesions.
 Method
o Washings: Specimen is collected by doctor in a clean `specimen trap’. Label
container with exact body site.
o Brushings: Insert the brush into a specimen container with 50% methanol /
ethanol or Surepath fixative or roll the contents of the brush onto a clean,
labelled glass slide and fix immediately with spray fixative.
 Specimen requirements:
o Amount of specimen: Washings: 10 ml preferred.
o Fixation: Equal volumes of 50% ethanol/methanol or Surepath fixative for
preservation can be added and may be stored in the fridge prior to processing.
do not add alcohol if Microbiology and Cytology is requested on one sample.
It is advisable to collect a second sample or send specimen to the laboratory
as soon as possible to be split.
o Transport: Bronchial washings should be transported to the laboratory
immediately.

BRONCHO-ALVEOLAR LAVAGE (BAL)

 Purpose: It is useful in the diagnosis of opportunistic infections in
immunocompromised hosts; it is also helpful to diagnose interstitial lung disease,
granulomatous disease and evaluation of transplant rejection. Malignancies and
certain other diseases can also be diagnosed.
 Method: Specimen is collected by doctor in a sterile, labelled container. BAL
specimens are obtained by wedging a sub segmental bronchus with the bronchoscope
and lavaging the area with saline or balanced salt solution.
 Specimen requirements:
o Amount of specimen: 10mlpreferred.
o Fixation: Equal volumes of 50% ethanol/methanol or Surepath fixative for
preservation and may be stored in the fridge prior to processing.Do not add
alcohol if Microbiology and Cytology is requested on one sample. It is advisable
to collect a second sample or send the specimen to the laboratory as soon as
possible, to be split.
o For Diff Count: Add equal amount of formalin and deliver immediately, on ice,
to cytology laboratory.
o Transport: Specimen should be transported immediately to the laboratory.

BODY FLUIDS – Peritoneal (ascites), Pleural, Pericardial, Cyst, etc.

When body fluids are tapped, they should always be submitted for cytological examination.
Large effusions, when initially tapped, may not be adequate due to degenerative changes or
a poor cellular yield. Cells lying in free fluid for too long a period of time often undergo severe
degenerative changes, which limit accurate cytology diagnosis.
If fluids need to be tapped repeatedly, re-accumulation of the fluids often yield more cells
which are better preserved. It is therefore recommended that each time an effusion is tapped,
that it is sent for cytological evaluation if a definitive diagnosis has not been made previously.
  Purpose: Body cavity fluids are usually collected for the diagnosis of malignant
neoplasms.
 Collection timing: Washings are usually obtained at the time of surgery.
 Method: Specimen is collected by doctor in a clean, properly labelled container.
 Specimen requirements:
o Amount of specimen: Minimum of 10ml – laboratory will accept up to 1000ml.
o Fixation: Equal volume of 50% ethanol/methanol or Surepath fixative for
preservation and may be stored in the fridge prior to processing.
o Transport: Specimens should be transported directly to the laboratory. Cells
remain interpretable in body fluids for several days after refrigeration.
. Collect 10-200 ml specimen in a leak-proof, screw-top, wide-mouth container. The
optimum sample size is 100 ml or larger. A cell block will be prepared if
adequate material is obtained. 


. 2 Immediately label the specimen container with the patient’s complete name and
source of specimen. 


. 3 Transport the specimen and requisition to the lab as soon as possible. If transport
is delayed by more than 10 minutes, add CytoLyt TM to the specimen (ratio of
50 percent specimen and 50 percent CytoLyt). 


. 4 If multiple tests are ordered on the same specimen (i.e., hematology, chemistry,
microbiology, etc.), divide the sample into appropriate tubes before adding
preservative. See the alphabetical test listing for specific sample requirements
for other tests. 


Body fluids (CSF, pleural effusion, peritoneal effusion, urine, cyst fluid etc.)

It is important that the specimens be sent to the laboratory as soon after collection as
possible. In some cases it is not possible to get the specimen down to the lab
immediately, but there are other things that can be done to preserve cellular detail.
If a biohazard fridge is available, refrigerate the specimen until delivering it to the
lab.
If cytolyt solution is available add an equal amount of cytolyt solution to the
specimen. Ex. 50mL urine add 50 mL of cytolyt solution.
If pre-filled cytojars are available they can also be used.

**If cytolyt is added please write "cytolyt" or "cytolyt added" to the container, as
processing these specimens differs from fresh specimens.
If a specimen is being split amongst various departments it is safer not to add cytolyt
because this can inhibit testing in certain instruments in other areas of the laboratory.
Inquire with the other laboratory departments if they can process specimens that are
fixed with cytolyt solution.
Fine needle aspirations/ Core biopsies (thyroid, lymph node, breast, cysts,etc)

Particularly for thyroid FNAs, on-site rapid assessment is recommended. Having a

cytotechnologist or pathologist there during the procedure ensures that an adequate
amount of material is collected. This keeps the patient from having to repeat a
procedure if the initial FNA was non diagnostic. If you are interested in on site rapid
assessment please contact me.

If performing a FNA without rapid assessment assistance here are some

recommendations:

If a biopsy is collected it is placed in 10% neutral buffered formalin. If one site is

being biopsied, then multiple cores can go in a single jar of formalin. If multiple sites
are being biopsied, each site needs its own individual jar. ex- head of pancreas
mass: three cores are taken they can all go into the same formalin jar. If there are
two lung nodules being biopsied (ex RLL and LUL) the cores from the RLL nodule
are to be placed in one formalin jar and the cores from the LUL nodule are placed in
another formalin jar. All jars need to be labeled appropriately.

If possible, before placing the core in formalin gently drag it across a glass slide and
allow it to air dry or place it in 95% alcohol. (whichever is easier)

If a FNA is being done and no core is obtained a drop of specimen is placed on a

glass slide and it is smeared with another slide via the direct smear method. (See
procedure on this page) This results in two slides containing specimen. One slide is
allowed to air dry and the other is placed in a slide holder containing 95% alcohol.
The rest of the specimen is put into a jar of cytolyt solution. As mentioned above, if
one site is being aspirated multiple passes of specimen can be placed in the same
cytolyt jar. If more than one site is being aspirated they must be collected separately.
Keep this in mind with the slides you prepare and make sure you label properly to
avoid confusion.

Efficacy of CytoLyt® hemolytic action on ThinPrep®

LBC using cultured osteosarcoma cell line LM8.
Norimatsu Y1, Ohsaki H, Masuno H, Kagawa A, Teramoto N, Kobayashi TK.
Author information
Abstract
OBJECTIVE:
The removal of blood components is necessary to improve the quality of the liquid-based
cytology (LBC) preparations. In ThinPrep® (TP) samples a cell suspension in a methanol-based
fixative undergoes a vacuum filtration method, whereas in SurePath™ (SP) samples a cell
suspension in an ethanol-based fixative is processed through a density gradient centrifugation
system prior to gravity deposition of the specimen onto a glass slide. We compared the cyto-
architectural features for the cytologic diagnosis of endometrial adenocarcinoma using parallel
TP and SP preparations in a previous publication.

STUDY DESIGN:
We performed our study on LM8 cells (a cultured osteosarcoma cell line). LM8 cells at a
concentration of 1.25 × 10(3) cell/cm(2) were seeded on a 35-mm plate in culture medium, which
contained 10% fetal bovine serum (FBS), 100 units/ml penicillin, and 100 μ/ml streptomycin in
Dulbecco's modified Eagle's medium (DMEM), and aliquots of the cell suspension obtained in
this way were compared after the addition of a hemolytic agent, i.e. Cytolyt® (CyL). LBC
preparations were then obtained on cell suspensions treated with CyL after different time
intervals of hemolysis.

RESULTS:
Treatment with CyL did not alter the cellularity of the preparation, but reduction of the nuclear
area and a tendency towards nuclear chromatin condensation with a subsequent higher
brightness were found. Because CyL is a 25% methanol-buffered solution, its alcoholic
concentration is low; it was our impression that, while its fixative effect was weak, its hemolytic
effect was high. Water influx or efflux through the cell membrane is controlled by osmotic
pressure changes induced by the buffer solution in the CyL solution. While CyL was not shown to
alter the cell shape, nuclear shrinkage was thought to be probably due to the increasing cell
dehydration caused by longer exposure intervals to methanol.

CONCLUSION:
This study has allowed us to make significant observations on the hemolytic properties of CyL,
and on its combined effects with PreservCyt on the cytomorphology of cells suspensions.