IG Participation Form

Laboratory Procedure for Immunoglobulin Gene Analysis

Contact and Shipping Details

First Name

Last Name

Institution (name that will appear on the diploma)

Address to send the samples

City

Country

Mobile number

Email Address

Verify Email Address

Additional Emails

If your country doesn't belong to the European Union, it is compulsory to provide the VAT/TAX number of your laboratory (HST, Federal Tax Number, ID...)

Tax Number

Patient Material

On completing the form, please tick the box that most accurately reflects your routine laboratory practice.

Requested material for accreditation

DNA (preferential)

Other on demand (viably frozen cells or RNA)

Other

Shipment

Regular mail - only DNA within Europe (paid by the organizers)

Transportation company (paid by the participant)

If shipment by transportation paid by the participant, please provide name of company

Source of cells

Peripheral Blood

Bone Marrow

Lymph Node

Other

Anticoagulant

EDTA Tubes

CPT Tubes

Heparinised Tubes

Other

Work-up of cells

Ficoll Gradient

Cell Suspension

Other

Nucleic Acid

DNA only

RNA only

Both

DNA Extraction Method

Please provide a brief description below

RNA Extraction / cDNA Synthesis Methods

Please provide a brief description below

PCR Methodology

General Strategy

Multiplex PCR

Single PCR

5´ Primer Used

IGHV Leader

IGHV FR1

Other

3´ Primer Used

Constant Region

IGHJ

Other

Reference for Primer Sets Used

Quantity of DNA / cDNA Used

Taq Polymerase Used (Including Company Name)

Evaluation / Processing of PCR Products

Analysis of PCR Products Prior to Sequencing

Yes

If yes, by

Agarose

PAGE

Genescan

Heteroduplex

Other

Purification of PCR Product Prior to Sequencing

Yes

Cloning of PCR Product Prior to Sequencing

Yes

If yes, please provide the situations and estimated frequency for when cloning is applied

Cloning Kit Used (including Company Name)

Prior to sequencing, do you check the clones for the correct insert?

Yes

Number of clones selected for Sequencing

Sequencing

General Strategy

Direct Sequencing

Cloning

Primers Used for Sequencing

Same as for PCR

Nested

M13

Other

Strands Sequenced

Forward only

Reverse only

Both

Bioinformatics Tools for Sequence Analysis - Please specify tools/programs used for: Visualisation of Chromatograms

Please specify tools/programs used for: Alignment of Both Strands (Forward and Reverse)

Please specify tools/programs used for: Multiple Sequence Alignment e.g. after Cloning

Please comment on any additional bioinformatic databases utilised

Bioinformatics Tools for Sequence Interpretation - Please specify the Tools/Programs utilised for each of the following: Identification of IGHV, IGHD, IGHJ Gene and Allele

Calculation of the Percentage Identity to the Germline

Do you exclude from the calculation the sequence corresponding to the 5´ primer?

Yes

Do you include in the calculation mutations within the HCDR3 of the IG gene?

Yes

Interpretation of Difficult Cases: Briefly outline how each of the following situations are handled/interpreted: Single Unproductive Rearrangements

Double Productive Rearrangements

Reporting IG Sequence Data in Clinical Routine Reporting IG Sequence Data in Clinical Routine

Briefly outline the information you currently provide in a Laboratory Report for IGHV Gene Mutation Status, e.g. tissue type, gDNA/cDNA, PCR primers, gene usage, % identity, functionality, conclusion, etc.

Briefly outline how you would report each of the following cases: 1. Mutated ´Borderline´ Rearrangement

2. Single Unproductive IGHV Rearrangement

3. Double Productive IGHV Rearrangements

4. Double Productive Rearrangements with Discordant Mutational Status

5. IGHV3-21 Rearrangement

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IG Participation Form

Requested material for accreditation

Shipment

If shipment by transportation paid by the participant, please provide name of company

Source of cells

Anticoagulant

Work-up of cells

Nucleic Acid

DNA Extraction Method

RNA Extraction / cDNA Synthesis Methods

General Strategy

5´ Primer Used

3´ Primer Used

Reference for Primer Sets Used

Quantity of DNA / cDNA Used

Taq Polymerase Used (Including Company Name)

Analysis of PCR Products Prior to Sequencing

If yes, by

Purification of PCR Product Prior to Sequencing

Cloning of PCR Product Prior to Sequencing

If yes, please provide the situations and estimated frequency for when cloning is applied

Cloning Kit Used (including Company Name)

Prior to sequencing, do you check the clones for the correct insert?

Number of clones selected for Sequencing

General Strategy

Primers Used for Sequencing

Strands Sequenced

Bioinformatics Tools for Sequence Analysis - Please specify tools/programs used for: Visualisation of Chromatograms

Please specify tools/programs used for: Alignment of Both Strands (Forward and Reverse)

Please specify tools/programs used for: Multiple Sequence Alignment e.g. after Cloning

Please comment on any additional bioinformatic databases utilised

Bioinformatics Tools for Sequence Interpretation - Please specify the Tools/Programs utilised for each of the following: Identification of IGHV, IGHD, IGHJ Gene and Allele

Calculation of the Percentage Identity to the Germline

Do you exclude from the calculation the sequence corresponding to the 5´ primer?

Do you include in the calculation mutations within the HCDR3 of the IG gene?

Interpretation of Difficult Cases: Briefly outline how each of the following situations are handled/interpreted: Single Unproductive Rearrangements

Double Productive Rearrangements

Briefly outline the information you currently provide in a Laboratory Report for IGHV Gene Mutation Status, e.g. tissue type, gDNA/cDNA, PCR primers, gene usage, % identity, functionality, conclusion, etc.

Briefly outline how you would report each of the following cases: 1. Mutated ´Borderline´ Rearrangement

2. Single Unproductive IGHV Rearrangement

3. Double Productive IGHV Rearrangements

4. Double Productive Rearrangements with Discordant Mutational Status

5. IGHV3-21 Rearrangement

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