Mohs/Histology Laboratory Compliance Manual...2 Quality Control Program It is the policy of Dr. laboratory to maintain a Quality Control Program to insure accuracy of results reported

Mohs/Histology

Laboratory Compliance Manual

Do Not Remove From Laboratory

INDEX

CLIA Quality Control Program page 2

Maintenance Record-Microscope page 6

Cryostat Protocol page 7

Maintenance Record-Cryostat page 8

Media or Stain Log page 9

Refrigerator Protocol page 10

Maintenance Record-Refrigerator page 11

Repair Companies page 14

Mohs page 16

Histopathology page 54

Job Descriptions page 90

1

Quality Assurance

The lab director will monitor all aspects of the laboratory and make certain that all testing

complies with the individual testing policy and procedure protocols.

Modifications and updates Date / Director Signature

Changes

2

Quality Control Program

It is the policy of Dr. laboratory to maintain a Quality Control

Program to insure accuracy of results reported. All employees of this laboratory must be

familiar with and adhere to all the policies herein stated with regard to quality control.

The Quality Control Program involves monitoring the facilities; test methods and

equipment, reagents, materials and supplies; procedure manual; method verification;

equipment maintenance; calibration and calibration verification; control procedures;

remedial actions; and maintenance of quality control records.

Facilities

The facilities of this laboratory consists of approximately square feet of

laboratory space. The environment in the working area of the laboratory will be

controlled by the commonly used heating, air-conditioning and ventilation equipment

used throughout the entire medical office, which will maintain a working temperature of

65 to 76 degrees Fahrenheit. Proper lighting will be monitored at least daily. Any burnt-

out bulbs will be replaced as soon as reasonably possible with new bulbs. The

Laboratory Director will ensure that proper ventilation systems are maintained

throughout the laboratory.

Test Methods, Equipment, Reagents, Materials and Supplies

The laboratory Director will be responsible for making the final decision on the test

system, equipment and methodologies used in the laboratory. The Laboratory Director

will ensure that proper test validation has been performed on all tests performed in the

laboratory before reporting any patient test results. The Laboratory Director will decide

the number of tests performed to validate the instrument or test system.

All equipment within the laboratory will be properly maintained according to the

manufacturer’s specifications. The Laboratory Director will be responsible for ensuring

that all maintenance and repair of equipment are completed in a timely fashion. All

maintenance and repairs will be recorded and maintained by the laboratory personnel and

will be reviewed by the Laboratory Director on a periodic basis.

The laboratory will maintain appropriate and sufficient supplies for the type and volume

of testing conducted by the laboratory. The Laboratory Director will be responsible for

maintaining an inventory of all supplies needed for the proper functioning of the

laboratory.

Temperature logs and graphs will be maintained for ambient room air and refrigerator

and freezer temperatures when necessary. Temperatures will be taken by an employee in

the laboratory and recorded. If a temperature is outside the desired range, the Laboratory

Director will be notified as soon as possible, and before any patient result is reported.

Appropriate corrective action will be taken and documented. It is the responsibility of

3

the Laboratory Director to determine the severity of the problem and the effect on the

testing process.

All reagents, solution, culture media, control materials, calibration materials and other

supplies will be labeled to indicate:

1) Identity and, when significant, titer, strength or concentration.

2) Recommended storage requirements.

3) Preparation and expiration dates.

4) Safety data.

5) Other pertinent information.

Reagents, solutions, culture media, controls, calibration materials and other supplies are

not used when they have exceeded their expiration dates, have deteriorated or are of

substandard quality.

Verification of Method Performance Specifications

The laboratory will establish performance specifications for any new test methodology

instituted after September 1, 1992. The laboratory will verify the performance

specifications for each new method based on:

1) Accuracy;

2) Precision;

3) Analytical sensitivity;

4) Analytical specificity to include interfering substances;

5) Reportable range of patient test results; and

6) Any other performance characteristic required to test performance.

The laboratory will document verification of applicable test performance specifications

and will maintain such documentation for the life of the methodology plus five years.

Equipment Maintenance

The Laboratory Director will ensure that periodic equipment maintenance and function

checks are performed as required by the manufacturer or determined by the Laboratory

Director. Proper records will be maintained to indicate the tests performed. These

4

records will be maintained for the life of the instrument plus five years. Maintenance

instructions for microscopes (see page 5) cryostats (see page 7), refrigerators (see page

10) should be documented.

Calibration and Calibration Verification

All instruments in the laboratory will be calibrated according to the following guidelines:

1) When a lot number of a reagent changes.

2) As recommended by the manufacturer.

3) When an “out of control” situation exists and cannot be resolved by other

means.

4) When major preventive maintenance, or replacement of critical parts

which may influence test performance, is done.

5) At least every six months.

Calibration verification will be performed according to the recommendations of the

manufacturer using calibration materials specified by the manufacturer.

Control Procedures

The control procedures set forth in the procedures manual will be followed for each

individual test.

Remedial Actions

Any remedial action taken by the laboratory will be documented on the Corrective Action

Request Form (see Corrective Action request Form, page 15) and reviewed by the

Laboratory Director. The records will be kept for a two-year period or until an inspection

occurs.

5

Equipment Quality Control for Microscopes

(State frequency of activity in spaces below)

1) Microscope stage and ocular eye pieces are to be cleaned

Stage is to be cleaned with alcohol or similar cleaner and ocular eye pieces

are to be cleaned with lens paper.

2) Grounding check is monitored

3) Notify the supervisor if any problems occur with instrumentation.

4) Every action is documented on the maintenance record form.

Protocol for Dealing with a Nonfunctioning Microscope

(Check applicable statement)

If, during an examination of specimens, the microscope should become nonfunctioning:

A back-up microscope is available at all times.

The test will be postponed until the microscope is required.

Other (Specify):

6

Maintenance Record for Microscopes

Date of maintenance activity must be recorded and initialed.

Stage & Oculars Cleaned Grounding/Cleaning

Date: Initial Date: Initial

7

Equipment Quality control for Cryostats

(State frequency of activity in spaces below)

1) Temperature is recorded daily and documented.

2) Temperature range is –20 degrees C to –30 degrees C.

3) Corrective action is taken and documented if temperature exceeds range.

4) Defrost of machine is done .

5) Interior is cleaned

using absolute alcohol, while wearing gloves.

6) Cryostat knives are sharpened as needed; or use disposable blades.

7) Air filter is cleaned as part of the maintenance every .

8) Thermometer check is done .

9) The fly wheel and moving components on the cryostat are oiled, as

recommended by the manufacturer, every .

10) Preventive maintenance and grounding check are done every

.

11) If any accidents occur while working with the cryostat, report to your

supervisor and the incident will be documented.

12) Notify supervisor if any problems occur with instrumentation.


Protocol for Dealing with a Nonfunctioning Cryostat

1) The cryostat is checked for proper functioning.

2) If the unit does not work, the patients who have been scheduled for

surgery on that day will be called immediately and surgery canceled.

3) The service contract company will then be called for repair. Repair or

replacement is guaranteed within 24 hours.

4) Instruction manuals are to located in the laboratory.

8

Maintenance Record for Cryostats

Month Year

Activity Clean Thermometer Moving Clean Preventive Defrost Problems/

Interior Check Components Air Maintenance Machine Supervisor

Filter Attention

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30

31

9

Media or Stain Receipt Log

Media or Manufacturer Batch Date

Stain Number Opened Condition Action taken if any

Notes:

10

Equipment Quality Control for Refrigerators

(State the frequency of activity in spaces below)

1) Read and record internal temperature daily. Optimum temperature for the

refrigerator is 2 degrees C to 8 degrees C (35.6 degrees F to 46.4 degrees

F) and for the freezer is –5 degrees C or colder (23 degrees F or colder).

2) Check door gasket seal every .

3) Defrost and clean interior every .

4) Grounding check is monitored every .

5) Notify the supervisor if any problems occur with instrumentation.


11

Temperature Monitor Log for Refrigerators, Freezers, Cryostats

Month Year

(Insert temperature in appropriate box)

Date Room Refrigerator Freezer Cryostat Initials

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31

12

Maintenance Record for Refrigerators

(Date of maintenance activity must be recorded and initialed.)

Check Door Gasket Seal Defrost/Clean Check Grounding Every six months Initial Every three months Initial Every three months Initial

13

Laboratory Quality Control for Hematoxylin and Eosin

Month Year

Nuclei Cytoplasm Thickness Section

Day Slide # Blue Pink/Red 5-10 Microns Complete Comments Initial

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31

Monthly Review By:

Date:

14

Repair Service Companies for Laboratory Equipment

(State Name, Address and Phone numbers of the repair service company in the spaces

below)

Microscope:

Cryostat:

Fume Hood:

Refrigerator:

15

Corrective Action Request Form

Quality Assurance Program

Corrective Action Request Form

Originator: Date:

Responsible Person:

Area of Concern:

Nature of Problem:

Signature

Corrective Action:

Signature

Reviewed By: Date:

Discussed at Staff Meeting Yes _____ No Date:

Comments:

Signature

16

Proficiency Testing

At the current time, there is no formal or approved procedure for proficiency testing

of Mohs’ surgical laboratories. There is discussion that the American College of

Mohs’ Micrographic Surgery and Oncology is developing such a program. When

that program is developed, this laboratory will be pleased to participate.

17

Laboratory Procedure

Histopathology – Mohs Surgery

Laboratory:

Director:

Because there may be some variation in the test procedures for histopathology –

Mohs surgery (e.g., stains used) you will need to thoroughly review the following

Laboratory Procedure Manual on histopathology – Mohs surgery and modify or

rewrite to conform with the practices in your laboratory. You may modify each

section in the space provided at the bottom of each page.

18

Record of Changes

All changes should be made in the space provided or on the corresponding facing

page. Each change or notation should be referenced to the appropriate paragraph

number and signed by the Laboratory Director.

Review Policy

This procedure manual is reviewed by the Laboratory Director annually and at other

times as required by major changes in procedure or other circumstances affecting

laboratory performance of the test.

Review by Laboratory Director

Date: Signature:

Date of first use of these procedures:

Date of last use of these procedures:

Copies of all procedures must be retained for two years after last date of use.

19



1) Principle of Test

2) Diagnostic Value

3) Specimen Requirements:

• Patient preparation

• Specimen collection

• Slide/specimen rejection

• Specimen handling, storage, preservation and identification

4) Materials and Reagents:

• Materials and reagents used

• Preparation and labeling

• Storage, use and handling

5) calibration

6) Quality Control

7) Test Procedure

8) Calculations

9) Reading and Reporting

• Reading results

• Reportable range

• Reference range

• Critical value reporting

• Procedures for panic values

20

• Reporting results

10) Procedure Notes

11) Limitations of Procedure

12) Remedial Actions

13) References

21


Reviewed by Director

1. Principle of Test

1.1 Provides microscopic data from tissue samples for correlation with

clinical data. Allows for precise margin control excision of

contiguously growing cutaneous neoplasms.

MODIFICATIONS AND UPDATES

Paragraph: Changes: Date: Director’s Signature

22

2. Diagnostic Value

2.1 Assessment of therapy, such as completeness of tumor removal.



23

3. Specimen Requirements

3.1 Patient Preparation

The patient is appropriately prepared for the type of surgical

procedure performed to obtain the specimen for histologic

examination.



24

3.2 SPECIMEN COLLECTON PROCEDURE

3.2.1 Mohs micrographic surgery

• Debulking of tumor

• Removal of a thin layer of tissue with a 1 – 3 mm margin

surrounding the tumor. A thin specimen allows the tissue to be

mounted so that the lateral and deep margins can be examined

in the same plane under the microscope after frozen section

processing.

• Tissue is precisely oriented on the patient prior to removal.

• The layer of tissue removed is sectioned and the edges of the

specimens are marked with colored dyes. The dyes can be

easily identified in frozen sections and permit accurate

orientation of excised tissue.

• A detailed map is made to correlate with the tissue removed.



25

3.3 SLIDE/SPECIMEN REJECTION

Incorrectly labeled specimen, e.g., wrong patient.

3.4 SPECIMEN HANDLING, STORAGE, PRESERVATION AND

IDENTIFICATION

3.4.1 Designate a small contamination area within the laboratory

to handle specimens.

3.4.2 Gloves must be worn to handle all specimens.

3.4.3 Mohs surgeon brings tissue to laboratory technician with

Mohs map and proper identification.

3.4.4 Mohs surgeon reviews specimen with laboratory

technician, noting site and instructing if there is a vertical

frozen section.



26

3.4.5 Laboratory technician notes time and logs it, with his/her

name on Mohs map.

3.4.6 Patient name, site, date, Mohs surgeon and laboratory

technician is written on Mohs log, along with total number

of slides processed.

3.4.7 If a vertical frozen section has been ordered by the Mohs

surgeon, this is written in the vertical frozen section log,

and a pathology sheet is filled out with patient name, site,

age, account number, date, Mohs surgeon and laboratory

technician.

3.4.8 Specimens are frozen in correct order in Cryostat.



27

3.4.9 Laboratory technician will cut frozen sections and stain.

Slides are labeled with number, full patient name, date and

type.

3.4.10 After cover slips are applied, the labeled slides with Mohs

map are brought to Mohs surgeon for evaluation.

3.4.11 Slides should be maintained for 10 years.

3.4.12 Specimens will be disposed of according to federal, state

and local law.



28

4. Materials and Reagents

4.1 MATERIALS AND REAGENTS USED

• Mounting compound or embedding medium

• Cryostat (specify)

• Histofreeze or comparable tissue freezing aerosol

• Glass slides or coated glass slides

• Staining materials (See pages 36 & 37)

• Dyes for marking tissue

• Cover glass



29

4.2 PREPARATION AND LABELING

4.2.1 Liquid reagents may be transferred to smaller containers for

that particular reagent

4.2.2 Dilutions of stock solutions should be performed under a

ventilation hood for volatile materials.

4.2.3 All reagents are to be labeled with the following

information:

• reagent

• dilution

• date prepared

• technician

• temperature for storage (if not room temperature)



30

4.3 STORAGE, USE AND HANDLING

(check below where appropriate)

4.3.1 Manufacturer’s control checks of media are not

provided; therefore, the laboratory will check each batch or

shipment of reagents when opened or prepared for positive and

negative reactivity as well as sterility. The laboratory will also

check media for their ability to support growth, and, as

appropriate, selectivity, inhibition and/or biochemical response.

All information will be properly documented.

Manufacturer’s control checks of media are provided and

the manufacturer’s product insert specifies that the manufacturer’s

quality control checks meet the National Committee for Clinical

Laboratory Standards for media quality control. The laboratory

will document the physical characteristics of each medium to

indicate that it is not compromised and report any deterioration to

the manufacturer. The laboratory will follow the manufacturer’s

specifications for using media. The laboratory will also check

each batch or shipment of reagents when opened or prepared for

positive and negative reactivity. All information will be properly

documented.



31

4.3.2 Reagents are stored according to manufacturer’s

instructions and temperature logs are maintained as

appropriate.

4.3.3 Do not use reagent after expiration date.

4.3.4 Discard and do not use unlabeled reagents.

4.3.5 Material safety data sheet are located

4.3.6 Reagents are disposed of according to federal, state, and

local laws.



32

5. Calibration

5.1.1 Temperature and time-controlled equipment is calibrated daily or

according to the manufacturer’s recommendations.

5.1.2 Daily temperature charts of all temperature-sensitive equipment

are maintained. Examples: refrigerators, freezers, tissue-

processing equipment, cryostat.

5.1.3 Ph meters are calibrated with each use.

5.1.4 While cutting frozen sections, the cryostat will be kept within a

range of –20 degrees C to –30 degrees C.



33

6. Quality Control

(Check where applicable)

6.1.1 All slides, paraffin blocks and reports are available for

review by laboratory accrediting agencies.

6.1.2 Reports from cases sent for consultation are maintained as

part of the record.

6.1.3 Reports of cases presented at conferences and scientific

meetings and in scientific publications are maintained as part of the

record.

6.1.4 The laboratory participates in the reciprocal reading of

slides with

6.1.5 A control slide will be made and evaluated each day that a

frozen section is prepared. A record of the control slide will be

maintained.



34

7. Test Procedure

7.1 PROCEDURE FOR SECTIONING THE SPECIMEN

• The specimens are brought into the laboratory by the Mohs

surgeon and given directly to the laboratory technician. The

tissue is presented with the epidermis, or the most superficial

side of the tissue, facing up.

• The tissue specimens are flattened and mounted on a cryostat

object disc using mounting compound or embedding medium.

The specimens are mounted so that the lateral and deep margins

will be in the same plane when sectioned.

• The cryostat object disc is placed in a cryostat for freezing.

Histofreeze or a comparable tissue freezing aerosol may be

used to expedite freezing.

• The tissue is then cut into 4 – 10 micron sections in a cryostat,

which is kept within a range of –20 degrees C to –30 degrees C.



35

• Sections are mounted on glass slides. These may be coated

with Poly-1-Lysine, egg albumin or other substances to help

ensure that the tissue adheres to the slides.

• After cutting, slides are handled according to the procedure

outlined under staining techniques.

• Do not touch other objects in rooms while wearing

contaminated gloves.

• Wipe outside surface of the cryostat with bleach.

• Wash hands after cutting frozen sections.

• All tissue disposed of are treated as biohazard waste according

to federal, state and local laws.



36

7.2 STAINING H AND E OF FROZEN SECTIONS

This procedure is to be used when sections are stained by hand and

not when an automated stainer is used.

7.2.1 Fix in acetone 100% 30 sec.

7.2.2 Drying time 20 sec.

7.2.3 Gill’s #3 hematoxylin * 60 sec.

7.2.4 Rinse in warm tap water until water runs clear

7.2.5 Eosin Y alcoholic ** 1 dip

7.2.6 100% ETOH 20 dips

7.2.7 100% ETOH 20 dips



37

7.2.8 100% ETOH 20 dips

7.2.9 Hisoclear *** 20 dips

7.2.10 Histoclear 20 dips

7.2.11 Histoclear 20 dips

7.2.12 Apply coverslip

* Hematoxylin (Harleco Brand) leaves least amount of

background residue

** Eosin Y

*** Histo-Clear (or any xylene substitute)

If the local tap water is not sufficient for bluing, the following may

be used:

Dip in acid alcohol for 15 seconds made by adding four

drops hydrochloric acid to container filled with 70%

alcohol. Rinse in tap water. Dip in ammonia water for 5

seconds made by adding four drops of ammonium

hydroxide to container filled with tap water. Rinse in

running tap water for 2 minutes. Proceed to Eosin.



38

7.3 STAINING H & E OF FROZEN SECTIONS

This procedure is to be used with the Shandon Linistain automated

stainer.

7.3.1 70% alcohol and formalin 30 sec.

7.3.2 Hematoxylin * 15 sec.

7.3.3 Hematoxylin * 15 sec.

7.3.4 Water wash 15 sec.


7.3.6 0.25% acid alcohol 15 sec.




39


7.3.9 Ammonia water 15 sec.


7.3.11 70% alcohol 15 sec.

7.3.12 Alcoholic Eosin 15 sec.

7.3.13 95% alcohol 15 sec.

7.3.14. 95% alcohol 30 sec.

7.3.15 100% alcohol 15 sec.



40

7.3.16 100% alcohol 30 sec.

7.3.17 Xylene substitute 30 sec.


* Harris’s modified hematoxylin which contains acetic acid.

** The acid alcohol is made by adding four drops hydrochloric

acid to the 15-second container filled with 70% alcohol.

*** The ammonia water is made by adding four drops of

ammonium hydroxide to the 15-second container filled

with tap water.

**** Eosin Y



41

7.4 STAINING TOLUIDINE BLUE OF FROZEN SECITONS

This is a procedure to be used with the Shandon Linistain

automated stainer.

7.4.1 Fix in absolute alcohol 30 sec.

7.4.2 Running water 60 sec.

7.4.3 Toluidine blue 0.8% aqueous * 30 sec.




42



7.4.7 95% alcohol 30 sec.

7.4.8 Absolute alcohol 30 sec.





43

7.4.11 Clearant (xylene substitute) 30 sec.



7.4.14 Clearant holding baths


* The Toluidine Blue is made by adding 0.8 grams Toluidine

Blue O Powder to 100 cc deionized water. Mix well. Filter before

using.



44

7.5 STAINING TOLUIDINE BLUE OF FROZEN SECTIONS

This procedure is to be used when sections are stained by hand and

not when an automated stainer is used.

7.5.1 Fix in absolute alcohol 30 sec.

7.5.2 Wash in water until all the mounting

media is dissolved, about 1 minute 60 sec.


7.5.4 Rinse in tap water to remove

excess stain. 15 sec.

7.5.5 95% alcohol 5 dips



45

7.5.6 95% alcohol 5 dips







* The Toluidine Blue is made by adding 0.8 grams Toluidine

Blue O Powder (Fisher Scientific) to 100 cc deionized water. Mix

well. Filter before using.



46

8. Calculations

Not applicable.



47

9. Reporting and Reading Results

9.1 READING RESULTS

9.1.1 Gross

Not applicable for Mohs Surgery

9.1.2 Microscopic

A detailed report should be recorded which includes a

description of the tumor and any unusual histologic features

as well as a description of where any residual tumor is

remaining after each layer of tissue is removed.

9.1.3 When performing Mohs surgery a map is used instead of a

report to decide where any residual tumor is remaining.

Notes are made on the map and/or is the record of any

unusual histologic features.



48

9.2 REPORTABLE RANGE

Not applicable

9.3 REFERENCE RANGE

Not applicable

9.4 CRITICAL VALUE REPORTING

Not applicable



49

9.5 PROCEDURES FOR PANIC VALUES

Not applicable

9.6 REPORTING RESULTS

The presence or absence of tumor and relationship to specimen

margins should be recorded. When performing Mohs surgery this

is recorded on the patient map.



50

10. Procedure Notes

Not applicable.



51

11. Limitations of Procedure

Mohs micrographic surgery is the most effective therapy for treating

cutaneous neoplasm that spread by contiguous growth, but it has limited

effectiveness in treating cutaneous neoplasms that grow in a

noncontiguous manner.



52

12. Remedial Actions

All out-of-control situations not resolved by a simple repeat analysis will

be reviewed by the Laboratory Director as soon as practical after the

event. The Laboratory Director will review the corrective action to assure

that appropriate action was taken and proper procedures were followed. A

Corrective Action Form will be filled out whenever a problem arises in

calibration or an out-of-control situation is not resolved by simple repeat

analysis.



53

13. Sources

Cottel, W. I., Bailin, P.L., et al. Essentials of Mohs Micrographic Surgery,

J. Dermatol, Surgery Oncol. 1988; 14:1, 11, - 13.

Mikhail, G. R.: Mohs Micrographic Surgery, W. B. Saunders Company,

1991.

Mohs, F. E., Chemosurgery: Microscopically Controlled Surgery for Skin

Cancer. Springfield, IL, Charles C. Thomas, 1978.

Roenigk, R. K., Mohs Micrographic Surgery, Mayo Clinic Proceedings

1988; 63:175-183.

Marsing Mohs lab consulting services 1999.

College of American Pathologists



54

Laboratory Procedure Manual – Histopathology

Laboratory:

Director:

Because there may be significant variations in the test procedures for

histopatholy, e.g., stains used, and because many dermatologists may send

specimens to an outside laboratory for preparation, you will need to

thoroughly review the following Laboratory Procedure Manual for

histopathology and modify or rewrite to conform with the practices in

your laboratory. You may modify each section in the space provided at

the bottom of each page.

55

RECORD OF CHANGES

All changes should be made in the space provided or on the corresponding

facing page. Each change or notation should be referenced to the

appropriate paragraph number and signed by the Laboratory Director.

REVIEW POLICY

This procedure manual is reviewed by the Laboratory Director annually

and at other times as required by major changes in procedure or other

circumstances affecting laboratory performance of the test.

REVIEW BY LABORATORY DIRECTOR

DATE: SIGNATURE:

Date of first use of these procedures

Date of last use of these procedures

Copies of all procedures must be retained for two years last date of use.

56

Histopathology



2. Diagnostic Value


• patient preparation

• specimen collection

• slide/specimen rejection

• specimen handling, storage, preservation and identification


• materials and reagents used

• preparation and labeling

• storage, use and handling

5. Calibration

6. Quality Control

7. Test Procedure

8. Calculations

9. Reading and Reporting

• reading results

• reportable range

• reference range

• critical value reporting

• procedures for panic values

57

• reporting results

10. Procedure Notes


12. Remedial Actions

13. Sources

58

Histopathology




clinical data.

1.2 Provides permanent data for concurrent and retrospective review.



59

2. Diagnostic Value


clinical data.



60


3.1 PATIENT PREPARATION

The patient is appropriately prepared for the type of surgical

procedure performed to obtain the specimen for histologic

examination.

3.2 SPECIMEN COLLECTION PROCEDURE

(Check appropriate surgical procedure)

Punch biopsy

Parallel incisional biopsy

Excisional biopsy



61

3.3 CRITERIA FOR SPECIMEN REJECTON

(Check applicable box)

3.3.1 Not applicable – specimen sent to laboratory for

specimen preparation. (Indicate laboratory name,

address, telephone number, contact person and

documentation of laboratory’s Quality Control

procedures).

OR

3.3.1 Unlabeled specimen container.

3.3.2 Incorrectly labeled specimen container, e.g., wrong

patient.

3.3.3 Specimens received in broken or crushed containers

can occasionally be salvaged, but the conditions of

receipt must be completely recorded.

3.3.4 Specimens received in the incorrect media or

fixative can occasionally be salvaged, but the

conditions of receipt must be completely recorded.



62

3.4 SPECIMEN HANDLING, STORAGE, PRESERVATION

AND IDENTIFICATION

3.4.1 Designate an area within the laboratory with good

ventilation for initial handling of the specimens.

3.4.2 Gloves must be worn to handle all specimens.

3.4.3 Assign specimen identification number and label all

worksheets, specimen containers and cassettes.

3.4.4 Record the patient’s name, date, specimen

identification number(s), anatomic sites and tests to

be performed.

3.4.5 Record gross description of specimen and

conditions of receipt.



63

3.4.6 Record description of sectioning of gross specimen.

3.4.7 Retain specimen containers until specimen is

processed and finalized.

3.4.8 Retain excess gross tissue in fixative for

(time), or longer if required by state law. Example:

retain the excess tissue if only a small representative

section was utilized for diagnosis.

3.4.9 Retain all slides and tissue blocks for 10 years or in

compliance with state law.

3.4.10 Specimens will be disposed of according to federal,

state and local laws.



64


4.1 MATERIALS AND REAGENTS USED


Not applicable – specimen sent for processing to: (Indicate

laboratory name, address, telephone number, contact person

and documentation of laboratory’s Quality Control

procedures).

OR

Disposable gloves

Laboratory aprons and lab coats

Cutting board

Forceps

Scalpel blade/knife/razor blades for gross sectioning

Tissue cassettes

Automated processor for fixation, dehydration and paraffin

embedding (specify)



65

Temperature-controlled paraffin embedding station (specify)

Microtome

Temperature-controlled water bath

Glass slides

Drying oven or microwave

Staining dishes or automated staining processor (specify)

Glass coverslips

10 percent buffered neutral formalin

Ethyl alcohol

Xylene (xylene substitutes may be used)

Paraffin

Staining reagents

Mounting media



66

4.2 PREPARATION AND LABELING

4.2.1 Liquid reagents may be transferred to smaller containers for

particular reagents.

4.2.2 Dilutions of stock solutions should be performed under a

ventilation hood for volatile materials.

4.2.3 All reagents are to be labeled with the following

information:

• Reagent

• Dilution

• Date prepared

• Technician

• Temperature for storage (if not room temperature)



67

4.3 STORAGE, USE AND HANDLING


4.3.1

Manufacturer’s control checks of media are not provided;

therefore, the laboratory will check each batch or shipment of

reagents when opened or prepared for positive and negative

reactivity as well as sterility. The laboratory will also check

media for their ability to support growth, and, as appropriate,

selectivity, inhibition and/or biochemical response. All

information will be properly documented.

Manufacturer’s control checks of media are provided and the

manufacturer’s product insert specifies that the manufacturer’s

quality control checks meet the National Committee for

Clinical Laboratory Standards for media quality control. The

laboratory will document the physical characteristics of each

medium to indicate that it is not compromised and report any

deterioration to the manufacturer. The laboratory will follow

the manufacturer’s specifications for using media. The

laboratory will also check each batch or shipment of reagents

when opened or prepared for positive and negative reactivity.

All information will be properly documented.



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4.3.2 Reagents are stored according to manufacturers’

instructions and temperature logs are maintained, as

appropriate.

4.3.3 Do not use reagent after expiration date.

4.3.4 Discard and do not use unlabeled reagents.

4.3.5 Material safety data sheets are located in MSDS section of

the manual.

4.3.6 Reagents are disposed of according to federal, state and

local laws.



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5. Calibration


5.1.1 Not applicable-specimens are sent for processing to:

(laboratory name, address, telephone number & contact person).

OR

5.1.1 Temperature and time-controlled equipment is calibrated

daily or according to the manufacturer’s recommendations.

5.1.2 Daily temperature charts of all temperature-sensitive

equipment are maintained. Examples: refrigerators,

freezers, tissue-processing equipment, embedding stations,

drying ovens, water baths.

5.1.3 ph meters are calibrated with each use.



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6. Quality Control


6.1.1 Not applicable-specimens are sent for processing to:

(laboratory name, address, telephone number & contact person).

OR

6.1.1 The laboratory participates in the Quality Control Program

administered by .

6.1.2 All slides, paraffin blocks and reports are available for

review by laboratory accrediting agencies.



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6.1.3 Reports from cases sent for consultation are maintained as

part of the record.

6.1.4 Reports of cases presented at conferences and scientific

meetings and in scientific publications are maintained as part of the

records.

6.1.5 The laboratory participates in the reciprocal reading of

slides with .

6.1.6 Other (specify)



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7. Test Procedure

(Check box below if applicable)

Not applicable – specimens are sent for processing to: (laboratory

name, address, telephone number & contact person).

OR

7.1 FIXATON

(Check applicable box below)

7.1.1 Tissue is usually fixed in 10% buffered neutral

formalin for 2 to 4 hours for small specimens (4x4x4 mm) and

up to 24 hours, depending upon size, for larger specimens.

7.1.2 Other fixatives may be used for special purposes

(specify).



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7.2 DEHYDRATION, CLEANING AND PARAFFIN

INFILTRATION

• 60 minutes 10% buffered neutral formalin

• 60 minutes 10% buffered neutral formalin

• 60 minutes 50% ethyl alcohol







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• 60 minutes xylene (xylene substitutes may be used)

• 60 minutes xylene (xylene substitutes may be used)

• 60 minutes paraffin at 60 degrees C

• 60 minutes paraffin at 60 degrees C



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7.5 TISSUE SECTIONING

7.4.1 Tissue sections are cut at 4 microns on the microtome.

7.4.2 Tissue sections are floated on a water bath at 45 degrees C

to 55 degrees C.

7.4.3 Tissue sections are picked up from the water bath onto

clean labeled slides.

7.4.4 Slides are dried in the drying oven for 10 minutes at 55

degrees C.



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7.5 STAINING

Mayer’s Hematoxylin and Eosin Procedure

Fixation: 10% buffered neutral formalin

Sections: Paraffin, 3 to 8 microns

Solutions:

Mayer’s Hematoxylin Stock Solution

Ammonium or Potassium alum 50.0 gm

Distilled water 1000 ml

Hematoxylin 1.0 gm

Sodium iodate 0.2 gm

Citric acid 1.0 gm

Chloral hydrate 50 gm



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Eosin Stock Solution

Eosin Y, water soluble 1.0 gm


Phloxine Stock Solution

Phloxine B 1.0 gm


Eosin-Phloxine working Solution

Combine in a 1000 ml cylinder:

Eosin stock solution 100 ml

Phloxine stock solution 10 ml

95% ethyl alcohol 780 ml

Glacial acetic acid 4 ml

Note: This solution is good for one week.



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7.5.1 Deparaffinize slides and hydrate to water

• 3 minutes xylene (xylene substitutes may be used:

specify )


specify )


specify )

• 1 minute 100% ethyl alcohol

• 1 minute 95% ethyl alcohol

• 1 minute distilled water

7.5.2 15 minutes Mayer’s hematoxylin solution

7.5.3 15 minutes wash in lukewarm running water

7.5.4 1 minute distilled water

7.5.5 1 minute 80% ethyl alcohol



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7.5.6 2 minutes eosin-phloxine solution

7.5.7 2 minutes 95% ethyl alcohol




7.5.11 2 minutes xylene

7.5.12 2 minutes xylene

7.5.13 Coverslip with mounting media



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8. Calculations

Not applicable



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9. Reading and Reporting Results

9.1 READING RESULTS

9.1.1 Gross

Size, color and shape will be recorded as part of the report.

In addition, a description of the gross sectioning will be

recorded. Diagrams may be used for clarity. Any unusual

aspects of receipt of the specimen should be noted.

Example: broken specimen bottle.

9.1.2 Microscopic

A detailed description of the microscopic features may be

recorded as part of the report. The results of control tissue

for the special stains should be recorded when appropriate.

9.1.3 Diagnosis

A final microscopic diagnosis should be recorded.



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9.2 REPORTABLE RANGE

All specimens require a report

9.3 REFERENCE RANGE

Not applicable



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9.4 CRITICAL VALUE REPORTING

Not applicable

9.5 PROCEDURES FOR PANIC VALUE

Not applicable



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9.6 REPORTING RESULTS

9.6.1 The final written report should contain the following

information:

• Patient name

• Unique number or demographic information pertaining

to patient

• Physician obtaining specimen

• Clinical information

• Anatomic site

• Clinical diagnosis or reason for test

• Microscopic diagnosis

• Physician making microscopic diagnosis

• Date of final report



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9.6.2 The following should be appended to the report as part of

the permanent record:

• Consultation reports

• Results of subsequent tests on the tissue

9.6.3 Distribution of reports

• Patient clinical record

• Laboratory file

9.6.4 Retention of reports

• The laboratory reports will be maintained for 10 years

or in compliance with local and state law.



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10. Procedure Notes

Not applicable



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The final microscopic diagnosis is highly dependent upon the submitted

tissue specimen.



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12 Remedial Actions

All out-of-control situations not resolved by a simple repeat analysis will

be reviewed by the Laboratory Director as soon as practical after the

event. The Laboratory Director will review the corrective action to assure

that appropriate action was taken and proper procedures were followed. A

Corrective Action Form will be filled out whenever a problem arises in

calibration or an out-of-control situation is not resolved by simple repeat

analysis.



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12. Sources

Farmer, E.R., Hood, A.F. (eds.): Pathology of the Skin. Appleton and

Lange, Norwalk, 1990

Prophet, E.B., Mills, B., Arrington, J.B., Sobin, L.G. (eds.): Laboratory

Methods in Histotechnology. American Registry of Pathology, 1992.

College of American Pathologists



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Job Descriptions for Laboratories

Performing Tests of High Complexity

LABORATORY DIRECTOR

The Laboratory Director is responsible for the overall operation and functioning

of the laboratory. The following represent the duties of a Laboratory Director:

1. Ensure that testing systems developed and used for each of the

tests performed in the laboratory provide quality laboratory

services for all aspects of test performance.

2. Ensure that the physical plant and environmental conditions of the

laboratory are appropriate for the testing performed and provide a

safe environment in which employees are protected from physical,

chemical and biological hazards.

3. Ensure that test methodologies selected are capable of providing

the quality of results required for patient care.

4. Ensure that the laboratory is enrolled in a HCFA-approved

proficiency testing program.

5. Ensure that quality control and quality assurance programs are

established and maintained to assure the quality of the laboratory

services provided and to identify failures in quality as they occur.

6. Ensure the establishment and maintenance of acceptable levels of

analytical performance of each test system.

7. Ensure that all necessary remedial actions are taken and

documented whenever significant deviations from the laboratory’s

established performance specifications are identified, and that

patient test results are reported only when the system is functioning

properly.

8. Ensure that reports of test results include pertinent information

required for interpretation.

9. Ensure that consultation is available to the laboratory’s clients on

matters relating to the quality of the test results reported and their

interpretation concerning specific patient conditions.

10. Employ a sufficient number of laboratory personnel with the

appropriate education and experience or training required to

provide appropriate consultation.

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11. Ensure that, prior to testing patients’ specimens, all personnel have

the appropriate education and experience, receive the appropriate

training for the type and complexity of the services offered, and

have demonstrated that they can perform all testing operations

reliably to provide and report accurate results.

12. Ensure that policies and procedures are established for monitoring

individuals who conduct preanalytical, analytical and

postanalytical phases of testing to assure that they are competent

and maintain their competency to process specimens, perform test

procedures and report test results promptly and proficiently. Also,

whenever necessary, identify the need for remedial training or

continuing education to improve skills.

13. Ensure that an approved procedure manual is available to all

personnel responsible for any aspect of the testing process.

14. Specify, in writing, the responsibilities and duties of each

consultant and each person engaged in the performance of the

preanalytic, analytic and postanalytic phases of testing, identifying

which examinations and procedures each individual is authorized

to perform.

Indicate below (Yes or No) whether the Laboratory Director will perform

this function.

Yes

No

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GENERAL SUPERVISOR

The General Supervisor is responsible for the day-to-day supervision of the

laboratory operation. The following represent the duties of a General Supervisor:

1. Responsible for providing day-to-day supervision of high

complexity test performance by testing personnel.

2. Must be on site to provide direct supervision when high

complexity testing is performed.

3. Responsible for monitoring test analyses and specimen

examinations to ensure that acceptable levels of analytic

performance are maintained.

4. Responsible for the annual evaluation and documentation of all

laboratory personnel.


this function.

Yes

No

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Job Description for Laboratories


TESTING PERSONNEL

The Testing Personnel are responsible for specimen processing, test performance

and for reporting test results. The following represent the duties of Testing

Personnel:

1. Follow the laboratory’s procedures for specimen handling and

processing, test analyses, and reporting and maintaining records of

patient test results.

2. Maintain records demonstrating that proficiency testing samples

are tested in the same manner as patient samples.

3. Adhere to the laboratory’s quality control policies. Also,

document all quality control activities, instrument and procedural

calibrations, and maintenance performed.

4. Follow the laboratory’s established corrective action policies and

procedures.

5. Be capable of identifying problems that may adversely affect test

performance or reporting of test results. Also, must be able to

either correct the problem or immediately notify appropriate

personnel.

6. Document all corrective actions taken when test systems deviate

from the laboratory’s established performance specifications.


this function.

Yes

No

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TECHNICAL CONSULTANT

The Technical Consultant is responsible for the technical and scientific oversight

of the laboratory. The following represent the duties of the Technical Consultant:

1. Select test methodology appropriate for the clinical use of the test

results.

2. Verify test procedures performed and establish laboratory’s test

performance characteristics, including the precision and accuracy

of each test and test system.

3. Enroll and participate in a HCFA-approved proficiency testing

program.

4. Establish a quality control program acceptable for the testing

performed in the laboratory.

5. Resolve technical problems and ensure that remedial actions are

taken whenever test systems deviate from the laboratory’s

established specifications.

6. Ensure that patient test results are not reported until corrective

action has been taken and the test system is functioning properly.

7. Evaluate the competency of all Testing Personnel and assure that

staff members maintain their competency to perform test

procedures and report test results promptly and accurately.

8. Evaluate and document the performance of individuals responsible

for high complexity testing at least semiannually during the first

year the individual tests patient specimens. Thereafter, evaluations

must be performed at least annually.


this function.

Yes

No

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CLINICAL CONSULTANT

The Clinical Consultant provides consultation regarding the appropriateness of

the testing ordered and interpretation of the test results. The following represent

the duties of the clinical Consultant:

1. Assist the laboratory’s clients in ensuring that appropriate tests are

ordered to meet the clinical expectations.

2. Ensure that reports of test results include pertinent information

required for specific patient interpretation.

3. Ensure that consultation is available and communicated to the

laboratory’s clients on matters related to the quality of the test

results reported and their interpretation concerning specific patient

conditions.

Documents

Mohs/Histology Laboratory Compliance Manual...2 Quality Control Program It is the policy of Dr. laboratory to maintain a Quality Control Program to insure accuracy of results reported